Methods for fixing carbon dioxide

Abstract

A system and method for fixing carbon dioxide by fermentation are provided. More specifically, the disclosure includes a step of producing methane by fermenting carbon dioxide with methane-producing archaea. The disclosure further provides integration of methane-producing fermentation with additional methods to achieve improved carbon efficiency.

Description

Technical Field

[0001] A method for fixing carbon dioxide is provided. More specifically, the method includes the step of fermenting carbon dioxide into methane and / or organic acids. The method can further include the step of cracking methane to produce hydrogen or hydrogen and CO for subsequent fermentation, or the step of directly providing methane to a methylotrophic fermentation process. Additional methods include the step of processing cell masses from fermentation into single cell proteins for use as nutritional supplements.

Background Art

[0002] Emissions of carbon monoxide and carbon dioxide from industrial processes are two major factors in climate change and global warming. Microbial fermentation can utilize microorganisms through their metabolic pathways to convert carbon monoxide (CO), hydrogen (H2), and / or carbon dioxide (CO2) into useful oxygenated hydrocarbon compounds such as ethanol, butanol, acetate, butyrate, 2,3-butanediol, and other desired products, thereby reducing such carbon emissions. Large-scale microbial fermentation also produces large amounts of microbial biomass. Conventionally, the disposal of microbial biomass requires extremely expensive waste treatment systems, storage sites, and landfill sites. Previous findings have shown that microbial biomass can be recovered and made into single cell protein (SCP) and other components and reused as a source of proteins, amino acids, and carbohydrates useful as nutritional supplements for animals, plants, or humans. For example, U.S. Patent No. 10,856,560 describes a method for producing whole cell animal feed by culturing acetogens to produce microbial biomass. Therefore, there is a need for methods and systems that can effectively convert carbon dioxide into products for other uses. Furthermore, there remains a need for methods and systems for efficiently converting microbial biomass into digestion-friendly nutritional supplements and compositions of any such nutritional supplements.

Summary of the Invention

[0003] In one embodiment, a method for converting CO2 includes the steps of fermenting a gaseous substrate containing CO2 and H2 with methane-producing archaea in a methane-producing microbial fermentation vessel, maintaining a CO2-to-H2 ratio of approximately 1:3 to approximately 1:4 in the gaseous substrate, and recovering methane from the methane-producing microbial fermentation vessel. In one embodiment, a method for converting CO2 includes the steps of: fermenting a gaseous substrate containing CO2 and H2 with methane-producing archaea in a methane-producing fermentation vessel; recovering methane from the methane-producing fermentation vessel; cracking at least a portion of the methane to produce H2; and returning at least a portion of the H2 to the methane-producing fermentation vessel. In another embodiment, a method for converting CO and CO2 includes the steps of: fermenting a gaseous substrate containing CO and H2 in a CO fermentation vessel with CO-converting acetic acid-producing bacteria to produce alcohol and CO2-containing vent gas; providing the CO2-containing vent gas from the CO fermentation vessel to a methane-producing bacteria fermentation vessel; and fermenting the CO2-containing vent gas in a methane-producing bacteria fermentation vessel with methane-producing archaea to produce methane.

[0004] In another embodiment, a method for converting CO and CO2 includes the steps of: fermenting a gaseous substrate containing CO and H2 in a CO fermentation vessel with CO-converting acetic acid-producing bacteria to produce alcohol and a first CO2-containing vent gas; and supplying the CO2-containing vent gas from the CO fermentation vessel to an acetic acid-producing CO2 fermentation vessel. The method further includes the steps of: fermenting the first CO2-containing vent gas in an acetic acid-producing CO2 fermentation vessel with CO2-converting acetic acid-producing bacteria to produce an organic acid and a second CO2-containing vent gas; supplying the organic acid to the CO fermentation vessel and supplying the second CO2-containing vent gas to a methane-producing bacteria fermentation vessel; and fermenting the second CO2-containing vent gas in a methane-producing bacteria fermentation vessel with methane-producing archaea to produce methane.

[0005] In another embodiment, a method for converting CO and CO2 includes the steps of: fermenting a gaseous substrate containing CO and H2 in a CO fermentation vessel with CO-converting acetic acid-producing bacteria to produce alcohol and a first CO2-containing vent gas; providing at least a portion of the first CO2-containing vent gas from the CO fermentation vessel to an acetic acid-producing CO2 fermentation vessel; fermenting at least a portion of the first CO2-containing vent gas in an acetic acid-producing CO2 fermentation vessel with CO2-converting acetic acid-producing bacteria to produce an organic acid; providing the organic acid to the CO fermentation vessel; providing at least another portion of the first CO2-containing vent gas from the CO fermentation vessel to a methane-producing bacteria fermentation vessel; and fermenting at least another portion of the first CO2-containing vent gas in a methane-producing bacteria fermentation vessel with methane-producing archaea to produce methane.

[0006] The above-mentioned features of this disclosure can be further described by reference to embodiments, some of which are illustrated in the accompanying drawings, so that the above-mentioned features of this disclosure may be understood in detail. However, it should be noted that the accompanying drawings only illustrate typical embodiments of this disclosure and should not be considered limiting in scope, for this disclosure may allow for other equally effective embodiments. [Brief explanation of the drawing]

[0007] [Figure 1] This figure shows a method for converting CO2, including methane cracking, which converts methane into hydrogen and solid carbon, and / or methane reforming, which produces hydrogen and CO. [Figure 2] This figure shows a method for converting CO and CO2, including fermentation using methanogenic archaea and fermentation using CO-converting acetic acid-producing bacteria. [Figure 3] This figure shows a method for converting CO and CO2, including fermentation using methanogenic archaea, fermentation using CO2-converting acetic acid-producing bacteria, and fermentation using CO-converting acetic acid-producing bacteria. [Modes for carrying out the invention]

[0008] The following description should not be taken as limiting, but is provided for the purpose of illustrating general principles of exemplary embodiments. The scope of this disclosure should be determined by reference to the claims. The term “approximately” modifying any quantity refers to the variation in that quantity encountered in real-world conditions, such as a laboratory, pilot plant, or production facility. For example, when a quantity of material or measurement used in a mixture or quantity is modified by “approximately,” it includes the variation and degree of care typically used when measuring under experimental conditions in a production plant or laboratory. For example, when a quantity of a product component is modified by “approximately,” it includes variation between batches of multiple experiments in a plant or laboratory and variation inherent in the analytical method. Whether modified by “approximately” or not, quantities include quantities equivalent to those quantities. Any quantity described herein and modified by “approximately” may also be used in this disclosure as an unmodified quantity. In the context of this disclosure, the use of the terms “a,” “an,” “the,” and similar demonstrative pronouns should be interpreted as encompassing both singular and plural forms unless otherwise indicated or more clearly negated in the context.

[0009] Unless otherwise indicated, the terms “comprising,” “including,” “having,” “containing,” or “characterized by” are inclusive and do not exclude any additional elements or method steps not described herein (i.e., “including but not limited to”). Any use of examples or illustrative language used herein (e.g., “such as,” “for example,” “for instance”) is intended solely to clarify the disclosure and, unless otherwise asserted, does not limit the scope of the disclosure.

[0010] Fermentation is a metabolic process used by microorganisms to generate energy for cell growth. Certain microorganisms can sustain their growth and produce oxygen-containing hydrocarbon compounds by fermenting C1-containing gaseous substrates, such as synthesis gas, carbon monoxide (CO)-containing gaseous substrates, or carbon dioxide (CO2)-containing gaseous substrates. In such cases, the microorganisms use one or more C1 components in the C1-containing gaseous substrate as the primary carbon source for their growth. Terms such as “fermentation,” “fermentation method,” and “microbial fermentation method” are intended to encompass both the growth phase and the product biosynthesis phase of the method. In anaerobic microbial fermentation methods, large quantities of microbial biomass are obtained, which can be purged out and processed into useful products such as nutrient supplements. Specifically, this disclosure includes a method for extracting nutrient supplements from microbial biomass by an anaerobic fermentation method. Fermentable gaseous substrates refer to C1-containing gaseous substrates that include one or more of CO, CO2, or CH2O2. Suitable gaseous substrates can include various synthesis gases (i.e., synthesis gas) and industrial off-gases.

[0011] Synthesis gas can be supplied from any known source. In one embodiment, synthesis gas can be supplied by gasification of carbonaceous material. Gasification involves partial combustion of biomass under conditions of limited oxygen supply. The resulting gas may contain CO, CO2, and H2. Several examples of suitable gasification methods and apparatus are described in U.S. Patent Applications 61 / 516,667, 61 / 516,704, and 61 / 516,646 (all filed April 6, 2011), and U.S. Patent Applications 13 / 427,144, 13 / 427,193, and 13 / 427,247 (all filed March 22, 2012), all of which are incorporated herein by reference. In another embodiment, synthesis gas can be produced from the electrolysis of water and carbon dioxide. In this embodiment, oxygen is removed from the obtained gas, and the obtained gas can be further blended with other gas sources to form a desired fermentable gaseous substrate.

[0012] Industrial off-gases may contain C1-containing waste gases from industrial processes that would normally be discharged into the atmosphere. Examples of industrial off-gases include gases produced during microbial fermentation, ferrous metal product manufacturing, non-ferrous product manufacturing, petroleum refining, coal gasification, electricity production, carbon black production, ammonia production, methanol production, coke production, and gas reforming. A C1-containing gaseous substrate may also contain H2. H2 can also be supplied to the C1-containing gaseous substrate separately to form a desired gas composition suitable for fermentation. Examples of H2 sources include gases produced during the manufacture of ferrous metal products, non-ferrous metal products, petroleum refining methods, coal gasification, biomass gasification, electricity production, carbon black production, ammonia production, methanol production, and coke production. Other hydrogen sources include, for example, H2O electrolysis and bio-generated H2. Fermentation of fermentable gaseous substrates using microorganisms is carried out in a fermentation vessel. The fermentation vessel includes a fermentation bioreactor consisting of one or more vessels and / or columns or piping, and may include batch reactors, semi-batch reactors, continuous reactors, continuous stirred tank reactors (CSTRs), bubble column reactors, external circulating loop reactors, internal circulating loop reactors, immobilized cell reactors (ICRs), trickle bed reactors (TBRs), mobile bed biofilm reactors (MBBRs), gas lift reactors, membrane reactors such as hollow fiber membrane bioreactors (HFMBRs), static mixers, gas lift fermenters, or other vessels or devices suitable for gas-liquid contact.

[0013] A culture medium suitable for anaerobic microbial growth and for fermenting a fermentable gaseous substrate into one or more oxygen-containing hydrocarbon compounds is added to the fermentation vessel to support the fermentation of the gaseous substrate by acetic acid-producing bacteria. Several examples of culture medium compositions are described in U.S. Patent Applications 16 / 530,502 and 16 / 530,481 filed August 2, 2019, and U.S. Patent No. 7,285,402 filed July 23, 2001, all of which are incorporated herein by reference. The culture medium can be sterilized to remove undesirable microorganisms and inoculate the fermentation vessel with desired microorganisms. Sterilization may not always be necessary. A culture medium suitable for methanogenic fermentation is described in U.S. Patent No. 11,401,499, which is incorporated herein by reference.

[0014] Methane production fermentation and methane cracking The method for converting CO2 illustrated in Figure 1 includes methane production in a methane-producing microbial fermentation vessel 105, and the use of a methane cracking device 120 to convert methane 135 into hydrogen 110 and solid carbon or carbon monoxide 130.

[0015] Methane productionIn one embodiment illustrated in Figure 1, the method includes a methane-producing microbial fermentation vessel 105 that can be integrated with an industrial process for producing CO2. In this embodiment, the methane-producing microbial fermentation vessel 105 contains a microbial culture capable of hydrogen-nutrient methane production (i.e., conversion of CO2 + H2 to methane). The methane-producing microbial fermentation vessel 105 is connected to a hydrogen source and a CO2 gas supply source 115. The hydrogen source may be a hydrogen-rich flow 110 produced by a methane cracking device 120. A separate hydrogen source may be provided to the methane-producing microbial fermentation vessel 105. H2 and CO2 may be added to the methane-producing microbial fermentation vessel 105 separately, or they may be blended together and then added to the methane-producing microbial fermentation vessel 105. The method includes the step of maintaining the CO2 to H2 ratio in the fermentation vessel 105 to approximately 1:5 to approximately 1:1, in another embodiment, approximately 1:5 to approximately 1:2, in another embodiment, approximately 1:5 to approximately 1:3, in another embodiment, approximately 1:5 to approximately 1:4, in another embodiment, approximately 1:4 to approximately 1:1, in another embodiment, approximately 1:4 to approximately 1:2, in another embodiment, approximately 1:4 to approximately 1:3, in another embodiment, approximately 1:3 to approximately 1:1, in another embodiment, approximately 1:3 to approximately 1:2, in another embodiment, approximately 1:2 to approximately 1:1. A total gas delivery rate in the range of approximately 0.2 to approximately 25 gas volume, in another embodiment, approximately 2 to approximately 16, in another embodiment, approximately 1 to 22, in another embodiment, approximately 0.5 to 20 (STP, standard temperature and pressure) / volume / min is preferred. The methanogenic fermentation vessel 105 can also produce a fermentation liquid broth 140 containing methanogenic archaea, which can be processed into single-cell proteins.

[0016] Suitable microbial cultures can be readily obtained from public collections of microorganisms or isolated from various environmental sources. Such sources include anaerobic soils and sands, swamps, wetlands, marshes, estuaries, dense algal mats, terrestrial and marine muds and sediments, deep-sea and deep wells, sewage and organic waste sites and treatment facilities, and animal intestines and feces. Many pure cultures of single species are preferred. Classified pure cultures are all members of the Archaea domain [Woese et al. Proc Natl Acad Sci USA 87:4576-4579 (1990) “Towards a natural system of organisms: Proposal for the domains Archaea, Bacteria, and Eucharya.”, incorporated herein by reference], and belong to four different classes of the Euryarchaea kingdom. Suitable organisms include four different genera within the class Methanobacteria (e.g., Methanobacterium alcaliphilum, Methanobacterium bryantii, Methanobacterium congolense, Methanobacterium defluvii, Methanobacterium espanolae, Methanobacterium formicicum, Methanobacterium ivanovii, Methanobacterium palustre, and Methanobacterium thermagregans). (thermaggregans), Methanobacterium uliginosum, Methanobrevibacter acididuransMethanobrevibacter acididurans, Methanobrevibacter arboriphilicus, Methanobrevibacter gottschalkii, Methanobrevibacter olleyae, Methanobrevibacter ruminantium, Methanobrevibacter smithii, Methanobrevibacter woesei, Methanobrevibacter wolinii, Methanothermobacter marburgensis, Methanothermobacter thermautotrophum Thermautotrophicum (also called Methanothermobacter thermoautotroiphicus), Methanothermobacter thermoflexus, Methanothermobacter thermophilus, Methanothermobacter wolfeii, Methanothermus sociabilis, and five different genera within the class Methanomicrobia (for example, Methanocorpusculum bavaricum, Methanocorpusculum parvum, Methanoculleus chikuoensis, and Methanoculleus submarinus) submarinus), Methanogenium frygidam(frigidum), Methanogenium liminatans, Methanogenium marinum, Methanosarcina acetivorans, Methanosarcina barkeri, Methanosarcina mazei, Methanosarcina thermophila, Methanomicrobium mobile), seven different genera within the class Methanococci (for example, Methanocaldococcus jannaschii, Methanococcus aeolicus, Methanococcus maripaldis) It was classified into several genera within the class Methanopyri (e.g., Methanocaldococcus maripaludis), Methanococcus vannielii, Methanococcus voltaei, Methanothermococcus thermolithotrophicus, Methanocaldococcus fervens, Methanocaldococcus indicus, Methanocaldococcus infernus, Methanocaldococcus vulcanius), and one genus within the class Methanopyri (e.g., Methanopyrus kandleri). Suitable cultures include those from public culture collections (e.g., the American Type Culture Collection, the Deutsche Sammlung von Mikroorganismen and Zellkulturen GmbH, and the Oregon)These are available from the Collection of Methanogens. Many suitable hydrogenotrophic methanogenic bacteria, isolated as pure cultures and available in public culture collections, have not yet been fully classified. Preferred pure culture organisms include Methanosarcinia barkeri, Methanococcus maripaldis, Methanothermobacter thermoautotrophicus, and Methanothermobacter marbulgensis.

[0017] Suitable cultures of mixtures of two or more microorganisms can also be readily isolated from designated environmental sources [Bryant et al. Archiv Microbiol 59:20-31 (1967) “Methanobacillus omelianskii, a symbiotic association of two species of bacteria.”, incorporated herein by reference]. The suitable mixture may be a consortium in which cells of two or more species are physically associated, or a vegetative symbiotic mixture in which two or more species cooperate metabolically without physical association. Mixed cultures may possess useful properties beyond those available from pure cultures of known hydrogenotrophic methanogenic bacteria. These properties include, for example, resistance to contaminants in the gas feed stream such as oxygen, ethanol, or other trace components, or aggregate growth that can increase the culture density and volumetric gas processing capacity of the culture.

[0018] A suitable culture of mixed organisms can also be obtained by combining cultures isolated from two or more sources. One or more species in the suitable mixed culture should be methanogenic archaea. Any non-archaeal species may be bacterial or eukaryotic. Suitable cultures can also be obtained by genetic modification of non-methanogenic organisms, in which genes essential for supporting hydrogenotrophic methanogenesis are transferred from methanogenic microorganisms or from combinations of microorganisms that may or may not be methanogenic by themselves. Suitable genetic modifications can be obtained by enzymatic or chemical synthesis of the necessary genes.

[0019] The methanogenic fermentation vessel 105 achieves continuous methane production using a continuous hydrogenotrophic methanogenesis culture operation under stable conditions. Examples of such suitable conditions are described in Schill, N., van Gulik, M., Voisard, D., & von Stockar, U. (1996) Biotcchnol & Biocng 51:645-658. “Continuous cultures limited by a gaseous substrate: development of a simple, unstructured mathematical model and experimental verification with Methanobacterium thermoautotrophicum”, which is incorporated herein by reference. The culture medium may consist of dilute mineral salts and should be adapted to the specific culture in use.

[0020] The concentrations of the various medium components for use in the methanogenic fermentation method are as follows.

Table 1

[0021] The medium in the methanogenic fermentation vessel 105 should be replenished at a rate suitable for maintaining useful concentrations of essential minerals and eliminating any metabolites that can inhibit methanogenesis. Dilution rates of less than 0.2 culture volumes / hour are preferred as they result in high volume concentrations of active methanogenic capacity. In one embodiment, the redox potential is maintained at less than -400mV or below during methane production. In another embodiment, the redox potential is maintained at less than -300mV or below, in yet another embodiment, at less than -200mV, and in yet another embodiment, at less than -100mV.

[0022] Alternatively, the culture temperature may be maintained near the optimal temperature for the growth of the organism used in the culture (for example, about 35°C to 37°C for mesophilic organisms such as Methanosarcinia barkeri and Methanococcus maripaldis, or about 60°C to 65°C for thermophilic bacteria such as Methanothermobacter thermoautotrophus, and about 85°C to 90°C for organisms such as Methanocardococcus janaskii, Methanocardococcus favens, Methanocardococcus indicus, Methanocardococcus infernus, and Methanocardococcus bulcanius). However, it is conceivable that temperatures above or below the optimal growth temperature may be used. In another embodiment, a reducing agent can be introduced into the fermentation process along with CO2 and hydrogen. The reducing agent can preferably be hydrogen sulfide or sodium sulfide. Hydrogen itself can be used as a reducing agent to maintain the oxidation-reduction potential of the culture within the range required for optimal performance of hydrogen nutrient methane production (<-100mV). Generally, hydrogen is provided at a concentration effective enough to convert at least a portion of the carbon dioxide in the bioreactor into methane. In another embodiment, the oxidation-reduction potential of the culture can be maintained at <-100mV via an electrochemical cell immersed in the culture medium.

[0023] In another embodiment, the method includes various methods and / or features for reducing the presence of oxygen in the CO2 stream fed to the bioreactor. When catalyzing methane formation using obligate anaerobic methanogenic archaea, the presence of oxygen can impair the performance of the method and contaminate the product gas. Therefore, reducing the presence of oxygen in the CO2 stream helps to improve the method. In one embodiment, the oxygen level is reduced by passing an H2 / CO2 mixed stream through a palladium catalyst before gas inflow into the fermentation vessel, converting any trace oxygen into water. In this embodiment, H2 is provided in an amount exceeding the amount required during culture at a ratio of 2:1 to the contaminating oxygen. In another embodiment, oxygen is removed by pretreatment of the gas stream in the bioreactor. In this embodiment, the reduced product can be provided by supplying a source of organic material that can serve as a substrate for oxidative fermentation (e.g., glucose, starch, cellulose, fermentation residue from an ethanol plant, whey residue, etc.). A microbial biological catalyst is selected to oxidative ferment the selected organic source and obtain CO2 from the contaminating oxygen. In this embodiment, additional H2 is supplied to enable the conversion of this additional CO2 to methane in the anaerobic fermenter.

[0024] The methane-producing microbial fermentation method yields a specific CO2 absorption of approximately 0.5 to 3 mmol of CO2 / min / cell per gram in another embodiment, approximately 1 to 2 mmol of CO2 / min / cell per gram in another embodiment, approximately 0.5 to 1 mmol of CO2 / min / cell per gram in another embodiment, approximately 1 to 3 mmol of CO2 / min / cell per gram in another embodiment, and approximately 0.5 to 2 mmol of CO2 / min / cell per gram in another embodiment. In this embodiment, the methane-producing microbial fermentation method is effective in yielding a CO2 conversion rate of 65% or more in another embodiment, 70% or more in another embodiment, 75% or more in another embodiment, 80% or more in another embodiment, 85% or more in another embodiment, 90% or more in another embodiment, 85% to 95% in another embodiment, and 90% to 99% in another embodiment. The method further yields cell densities of up to 100 g / L, in one embodiment 10-80 g / L, in one embodiment 15-60 g / L, in one embodiment 20-50 g / L, in one embodiment 10-30 g / L, and in another embodiment 15-45 g / L.

[0025] The method further yields a specific H2 absorption of approximately 3 to 12 mmol of H2 / min / cell per gram, in another embodiment, approximately 3 to 10 mmol of H2 / min / cell per gram, in another embodiment, approximately 3 to 8 mmol of H2 / min / cell per gram, in another embodiment, approximately 3 to 6 mmol of H2 / min / cell per gram, in another embodiment, approximately 4 to 12 mmol of H2 / min / cell per gram, in another embodiment, approximately 4 to 10 mmol of H2 / min / cell per gram, in another embodiment, approximately 4 to 8 mmol of H2 / min / cell per gram, in another embodiment, approximately 5 to 12 mmol of H2 / min / cell per gram, in another embodiment, approximately 5 to 10 mmol of H2 / min / cell per gram, and in another embodiment, approximately 5 to 8 mmol of H2 / min / cell per gram. The method further provides cell retention times of approximately 5 to 50 hours, in another embodiment, approximately 5 to 40 hours, in another embodiment, approximately 5 to 30 hours, in another embodiment, approximately 5 to 25 hours, in another embodiment, approximately 5 to 20 hours, in another embodiment, approximately 5 to 10 hours, in another embodiment, approximately 5 to 8 hours, in another embodiment, and approximately 8 to 15 hours.

[0026] The method further yields methane production of approximately 0.4 to approximately 3 mmol of methane / min / cell per gram, in another embodiment, approximately 0.4 to approximately 2 mmol of methane / min / cell per gram, in another embodiment, approximately 0.4 to approximately 1 mmol of methane / min / cell per gram, in another embodiment, approximately 1.0 to approximately 3 mmol of methane / min / cell per gram, in another embodiment, approximately 1.0 to approximately 2.5 mmol of methane / min / cell per gram, in another embodiment, approximately 1.0 to approximately 2.5 mmol of methane / min / cell per gram, and in another embodiment, approximately 1.5 to approximately 2.5 mmol of methane / min / cell per gram. In this embodiment, the method is effective in producing a methane-effluent gas having a methane concentration greater than 60%, in another embodiment greater than 65%, in another embodiment greater than 70%, in another embodiment greater than 75%, in another embodiment greater than 80%, and in yet another embodiment greater than 85%.

[0027] Metanic Cracking As further illustrated in Figure 1, the methane 135 produced by the fermentation vessel 105 can be transported via line 136 to the methane cracking unit 120, or to drive various processes, and / or stored and sold as fuel (all referred to as 137). The methane cracking unit 120 can pyrolyze the methane into hydrogen 110 and solid carbon 130, or reform the methane into hydrogen 110 and carbon monoxide 130. In this specification, methane cracking can include both pyrolysis and reforming. Methane pyrolysis can be achieved by known methane pyrolysis methods, including, for example, microwave pyrolysis, molten metal pyrolysis, plasma arc pyrolysis, and combinations thereof. Examples of methane reforming include steam reforming, dry reforming, and partial oxidation.

[0028] In one embodiment, methane pyrolysis is carried out using microwave pyrolysis. Microwave pyrolysis is described in International Publication No. 2022 / 232942, published November 10, 2022, which is incorporated herein by reference. The method comprises the steps of providing a methane-containing feedstock to a microwave-inert reaction vessel or to a reaction vessel inert to both microwaves and radio waves. Solid carbon is present inside the vessel, and molecular oxygen is absent or present in negligible amounts. Water and molecular oxygen are not added to the vessel. The sole source of water and molecular oxygen inside the reaction vessel should be the small amounts of water, molecular oxygen, and oxygen-containing species (CO2) present in the feedstock. The solid carbon is then exposed to microwaves, radio waves, or both microwaves and radio waves, resulting in the carbon being present at a temperature of at least 200 Kelvin. This hot carbon heats the gaseous hydrocarbons, thereby forming hydrogen and additional solid carbon. Hydrogen and solid carbon are separated, and the hydrogen 110 can be returned to the methane-producing microbial fermentation vessel 105. In another embodiment, methane pyrolysis is carried out using molten metal pyrolysis. In yet another embodiment, methane pyrolysis is carried out using plasma arc pyrolysis.

[0029] CO fermentation and methane production fermentation A method for converting CO and CO2 as illustrated in Figure 2 includes the step of providing a CO-containing gaseous substrate 50 to a CO fermentation vessel 230. CO-converting acetic acid-producing bacteria in the CO fermentation vessel 230 can convert CO into one or more alcohols 255. Vent gas 481 from the CO fermentation vessel 230 contains CO2. Vent gas 481 may be supplied to a methane-producing fermentation vessel 105. A gaseous substrate 112 containing H2 and / or additional CO2 may also be supplied to the methane-producing fermentation vessel 105. Methane-producing archaea in the methane-producing fermentation vessel convert CO2 into methane 135. The methane 135 may be sold as "green methane / renewable natural gas" or sent to methane cracking. In another embodiment, H2 and / or CO from methane cracking may be supplied to the methane-producing fermentation vessel and / or CO fermentation vessel. Clostridium fermentation liquid broth 270 and / or methanogenic fermentation liquid broth 140 can be processed into single-cell proteins.

[0030] Methane production In one embodiment illustrated in Figure 2, the method includes a methane-producing microbial fermentation vessel 105 as described herein in the description of Figure 1. Metanic Cracking In any embodiment not shown in Figure 2, a portion of the methane 135 produced in the methane-producing microbial fermentation vessel 105 may be supplied to a methane-cracking apparatus as described herein in relation to Figure 1. The methane-cracking apparatus may be operated to yield a gaseous substrate containing H2 and / or CO. In one embodiment, the gaseous substrate containing H2 and CO may be separated into an H2-rich stream and a CO-rich synthesis gas. The H2-rich stream may be supplied to the methane-producing microbial fermentation vessel 105, and the CO-rich synthesis gas may be supplied to the CO-fermentation vessel 230. In another embodiment, the gaseous substrate containing H2 and CO may be supplied directly to the CO-fermentation vessel 230. In yet another embodiment, CO is not produced from the methane-cracking apparatus. In this scenario, the H2-containing gaseous substrate produced from the methane-cracking apparatus may be supplied directly to the methane-producing microbial fermentation vessel 105.

[0031] CO fermentation Certain types of acetic acid-producing bacteria can ferment the CO-containing gaseous substrate 50 in the CO fermentation vessel 230 into useful oxygen-containing hydrocarbon compounds 255 such as ethanol and butanol, thereby producing a fermented liquid broth containing acetic acid-producing bacteria 270. In this embodiment, the preferred gaseous substrate 50 contains at least about 5 mol% CO, in one embodiment at least about 10 mol% CO, in one embodiment at least about 20 mol% CO, in one embodiment at least about 30 mol% CO, in one embodiment about 10 to about 100 mol% CO, in another embodiment about 20 to about 100 mol% CO, in another embodiment about 30 to about 90 mol% CO, in another embodiment about 40 to about 80 mol% CO, and in another embodiment about 50 to about 70 mol% CO. In this embodiment, the CO-containing gaseous substrate 50 may have about 40 mol% or less of CO2; in one embodiment, the CO-containing gaseous substrate 50 may have about 30 mol% or less of CO2; in one embodiment, the CO-containing gaseous substrate 50 may have about 20 mol% or less of CO2; in another embodiment, the CO-containing gaseous substrate 50 may have about 10 mol% or less of CO2; in yet another embodiment, the CO-containing gaseous substrate 50 may have about 1 mol% or less of CO2; and in yet another embodiment, the CO-containing gaseous substrate 50 may contain no CO2 or substantially no CO2.

[0032] Depending on the composition of the CO-containing gaseous substrate 50, the CO-containing gaseous substrate 50 may be supplied directly to the fermentation vessel 230, or it may be further modified or blended to have an appropriate H2-to-CO molar ratio. In one embodiment, the CO-containing gaseous substrate supplied to the fermentation vessel has an H2-to-CO molar ratio of about 0.1 or more, in another embodiment, about 0.2 or more, in another embodiment, about 0.25 or more, and in another embodiment, about 0.5 or more. In one embodiment, H2 and / or CO from a methane cracking device may be supplied to the CO fermentation vessel 230.

[0033] The concentrations of various culture medium components used in the CO bioconversion fermentation method are as follows: [Table 2]

[0034] Examples of acetic acid-producing bacteria useful in CO bioconversion fermentation methods include Blautia producta, Butyribacterium methylotrophicum, Caldanaerobacter subterraneous, Caldanaerobacter subterraneous pacificus, Carboxydothermus hydrogenoformans, Clostridium aceticum, Clostridium acetobutylicum, Clostridium acetobutylicum P262, and Clostridium autoethanogenum (DSM). Clostridium autoethanogenum (DSM 10061, DSMZ), Clostridium autoethanogenum (DSM 23693, DSMZ), Clostridium autoethanogenum (DSM 24138, DSMZ), Clostridium carboxidivorans, Clostridium coskatii (ATCC PTA-10522), Clostridium drakei, Clostridium ljungdahlii PETC (ATCC) 49587), Clostridium ljungdahlii ERI2 (ATCC 55380), Clostridium ljungdahlii C-01 (ATCCClostridium ljungdahlii O-52 (ATCC 55889), Clostridium magnum, Clostridium pasteurianum (DSM 525, German DSMZ), Clostridium ragsdalei P11 (ATCC BAA-622), Clostridium scatologenes, Clostridium thermoaceticum, Clostridium ultunense, Desulfotomaculum kuznetsovii, Eubacterium limosam Examples include *Lyophyllum limosum*, *Geobacter sulfurreducens*, *Methanosarcina acetivorans*, *Methanosarcina barkeri*, *Oxobacter pfennigii*, *Peptostreptococcus productus*, *Clostridium stick-landii*, and mixtures thereof.

[0035] Anaerobic bacteria are bacteria that do not require oxygen for growth. If oxygen is present above a certain threshold, anaerobic bacteria may react negatively and even die. Acetate-producing bacteria are microorganisms that can produce acetate under anaerobic respiration or fermentation by utilizing the Wood-Ljungdahl pathway as their primary mechanism for energy conservation. Other useful oxygen-containing hydrocarbon compounds, such as formic acid, propionic acid, butyric acid, heptanoic acid, decanoic acid, ethanol, butanol, 2-butanol, and 2,3-butanediol, can also be produced by acetic acid-producing bacteria. Examples of acetic acid-producing bacteria suitable for converting C1-containing gaseous substrates into useful oxygen-containing hydrocarbon compounds include strains of Clostridium lyngdaryi, including those described in International Publication No. 2000 / 68407, European Patent No. 117309, U.S. Patents No. 5,173,429, 5,593,886 and 6,368,819, International Publication No. 1998 / 00558 and 2002 / 08438, and strains of Clostridium autoethanogenum (DSM 10061 and DSM 19630, DSMZ, Germany), including those described in International Publication No. 2007 / 117157 and 2009 / 151342, U.S. Patent No. 7,704,723 and “Biofuels and Bioproducts from Biomass-Generated Synthesis Gas”, Hasan Atiyeh, presented in Examples of Clostridium bacteria include Clostridium lagsudarei (P11, ATCC BAA-622) and Alkalibaculum bacchi (CP11, ATCC BAA-1772), as well as Clostridium carboxydivorans (ATCC PTA-7827), as described in U.S. Patent Application No. 2007 / 0276447.Other suitable microorganisms include those of the genus Moorella, including the Moorella species HUC22-1, and those of the genus Carboxydothermus. Each of these references is incorporated herein by reference.

[0036] CO fermentation can be carried out under reaction conditions appropriate to the desired fermentation mode. For example, in one embodiment, CO fermentation can be set up in a mode focused on CO-to-oxygenated hydrocarbon compound (e.g., ethanol) production. In this mode, approximately 4% to 6% of the carbon derived from the CO fed into CO fermentation is converted into biomass. In another embodiment, CO fermentation can be set up in a mode focused on CO-to-microbial biomass production. In this mode, approximately 6% to 7.5% of the carbon derived from the CO fed into CO fermentation is converted into biomass. Reaction conditions to be considered include pressure, temperature, gas flow rate, liquid flow rate, medium pH, oxidation-reduction potential of the medium, stirring speed (if using a stirred-tank reactor), inoculation level, appropriate gas substrate concentration to ensure that the CO in the liquid phase is neither limiting nor inhibiting, and appropriate product concentration to avoid product inhibition. The CO fermentation method yields CO conversion rates of 80% or more in one embodiment, 85% or more in another embodiment, 90% or more in yet another embodiment, 80% to 99% in yet another embodiment, 85% to 98% in yet another embodiment, and 90% to 97% in yet another embodiment. During the CO fermentation method, a cell density of 5 g / L or more is maintained in one embodiment, 10 g / L or more in one embodiment, 12 g / L or more in one embodiment, 8 to 15 g / L in another embodiment, 10 to 20 g / L in another embodiment, 12 to 25 g / L in yet another embodiment, and 10 to 30 g / L in yet another embodiment.

[0037] CO fermentation further yields a specific alcohol production capacity of approximately 10 grams or more of alcohol / day / cell in grams, in another embodiment, a specific alcohol production rate of approximately 12 g or more / day / cell in grams, in another embodiment, a specific alcohol production rate of approximately 14 g or more / day / cell in grams, in another embodiment, a specific alcohol production rate of approximately 10 to approximately 16 g / day / cell in grams, in another embodiment, approximately 10 to approximately 14 g / day / cell in grams, in another embodiment, approximately 10 to approximately 12 g / day / cell in grams, in another embodiment, approximately 10 to approximately 16 g / day / cell in grams, in another embodiment, approximately 10 to approximately 14 g / day / cell in grams, in another embodiment, approximately 12 to approximately 16 g / day / cell in grams, and in another embodiment, approximately 12 to approximately 14 g / day / cell in grams. CO-converting acetic acid-producing bacteria convert CO to produce one or more alcohols and a CO2-containing vent gas. The CO2-containing vent gas is then sent to a methane-producing fermentation vessel. The CO2-containing vent gas contains 5% or less CO, in one embodiment, 3% or less CO, in one embodiment, 2% or less CO, and in another embodiment, 1% or less CO. CO can be removed from the CO2-containing vent gas before it enters the methane-producing fermentation vessel to avoid CO inhibition.

[0038] CO fermentation, acetic acid production CO 2 Fermentation and methane production fermentation The method for converting CO and CO2 illustrated in Figure 3 includes the step of fermenting a gaseous substrate 50 containing CO in a CO fermentation vessel 230. CO-converting acetic acid-producing bacteria in the CO fermentation vessel 230 convert CO into one or more alcohols 255. The vent gas 481 from the CO fermentation vessel 230 contains CO2. 。Vent gas 481 can be supplied to the acetic acid production CO2 fermentation vessel 220 and / or the methane production fermentation vessel 105. The acetic acid production CO2 fermentation vessel 220 produces organic acids 227, which can be supplied to the CO fermentation vessel 230 to increase productivity in CO fermentation by producing oxygen-containing hydrocarbon compounds 255 such as ethanol and butanol. The vent gas 225 from the acetic acid production CO2 fermentation vessel 220 may contain unconverted CO2 and / or H2, which can then be delivered to the methane production fermentation vessel 105. The CO fermentation broth 270, the acetic acid production CO2 fermentation broth 250, and / or the methane-producing fermentation broth 140 can be further purged out of the fermentation vessels and processed into single-cell protein nutritional supplements.

[0039] Methane production In one embodiment illustrated in Figure 2, the method includes a methane-producing microbial fermentation vessel 105 as described herein in the description of Figure 1. In this embodiment, the methane-producing microbial fermentation vessel 105 can be used to maintain a balance between the organic acid consumption of the CO fermentation vessel 230 and the CO2 consumption of the acetic acid-producing CO2 fermentation vessel 220. A gaseous substrate 112 containing supplemental H2 and / or external CO2 may also be provided to the methane-producing microbial fermentation vessel 105 and / or the acetic acid-producing CO2 fermentation vessel 220.

[0040] CO 2 Acetate-producing bacteria and acetic acid-producing CO 2 fermentationCO2-converting acetic acid-producing bacteria can ferment a CO2-containing gaseous substrate into useful oxygen-containing hydrocarbon compounds such as C1-C10 organic acids 227, examples of which include acetic acid and butyric acid. In this embodiment, the acetic acid-producing CO2 fermentation vessel may be supplied with supplemental H2112. In one embodiment, the organic acid 227 may be supplied to the CO fermentation vessel 230, and the vent gas 225 may be delivered to the methane-producing bacteria fermentation vessel 105. In one embodiment, a suitable CO2-containing gaseous substrate contains at least about 10 mol% CO2, in one embodiment, at least about 20 mol%, in one embodiment, at least about 30 mol%, in one embodiment, at least about 40 mol%, in one embodiment, about 10 to about 70 mol%, in another embodiment, about 20 to about 70 mol% CO2, in another embodiment, about 30 to about 70 mol% CO2, in another embodiment, about 40 to about 70 mol% CO2, in another embodiment, about 10 to about 50 mol% CO2, in another embodiment, about 20 to about 40 mol% CO2, and in yet another embodiment, about 30 to 50 mol% CO2. In one embodiment, the CO2-containing gaseous substrate contains about 50 mol% or less of CO; in one embodiment, the CO2-containing gaseous substrate contains about 40 mol% or less of CO; in one embodiment, the CO2-containing gaseous substrate contains about 30 mol% or less of CO; in one embodiment, the CO2-containing gaseous substrate contains about 20 mol% or less of CO; in one embodiment, the CO2-containing gaseous substrate contains about 10 mol% or less of CO; in one embodiment, the CO2-containing gaseous substrate contains about 5 mol% or less of CO; in one embodiment, the CO2-containing gaseous substrate contains about 1 mol% or less of CO; and in another embodiment, the CO2-containing gaseous substrate contains no CO or substantially no CO.

[0041] Depending on the composition of the CO2-containing gaseous substrate, it may be directly supplied to the acetic acid production CO2 fermentation method, or it may be further modified or blended to contain an appropriate H2-to-CO2 molar ratio. For example, a stream containing high concentrations of CO2, such as exhaust gas from an industrial process, can be combined with a stream containing high concentrations of H2, such as off-gas from a coke oven. In one embodiment, the gaseous substrate supplied to the fermentation vessel has an H2-to-CO2 molar ratio of about 4:1 to about 1:2, in another embodiment, about 4:1 to about 1:1, in yet another embodiment, about 4:1 to about 2:1, and in yet another embodiment, about 3.5:1 to about 1.5:1.

[0042] The concentrations of various culture medium components used in the CO2 bioconversion fermentation method are as follows: [Table 3]

[0043] Acetogenic bacteria suitable for CO2 bioconversion contain a sodium pump, which may also be described as a sodium-transporting ATPase (in the case of membrane bioenergetics). Sodium-transporting ATPases are described in Muller, “Energy Conservation in Acetogenic Bacteria,” Appl. Environ. Microbiol. November 2003, vol. 69, no. 11, pp. 6345-6353, which is incorporated herein by reference. Acetogenic bacteria containing sodium-transporting ATPases require approximately 500 ppm of NaCl in their growth medium for growth. To determine whether acetic acid bacteria contain sodium-transporting ATPases, acetogens are inoculated into serum bottles containing approximately 30-50 ml of growth medium with approximately 0-2000 ppm of NaCl. Normal growth at NaCl concentrations above approximately 500 ppm indicates that the acetic acid bacteria contain sodium-transporting ATPases. In this embodiment, the composition of the fermentation medium includes a sodium ion concentration of about 40 to about 500 mmol / liter, in another embodiment, a sodium ion concentration of about 40 to about 250 mmol / liter, and in another embodiment, a sodium ion concentration of about 50 to about 200 mmol / liter. In one embodiment, the sodium ion concentration is about 500 ppm to about 8000 ppm, in another embodiment, about 1000 ppm to about 7000 ppm, in another embodiment, about 3000 ppm to about 6000 ppm, in another embodiment, about 2000 to about 5000 ppm, and in another embodiment, about 3000 to about 4000 ppm.

[0044] Examples of CO2-converting acetic acid-producing bacteria useful for CO2 bioconversion include Thermoanaerobacter kivui, Acetoanaerobium noterae, Acetobacterium woodii, Alkalibacrum batchii CP11 (ATCC BAA-1772), Moorella thermoacetica, Moorella thermoautotrophica, Ruminococcus productus, Thermoanaerobacter kivui, and combinations thereof.

[0045] Acetate-producing bacteria for CO2 fermentation can produce C1-C10 organic acids. In one embodiment, the organic acid is acetic acid. In another embodiment, the organic acid is butyric acid. In yet another embodiment, the organic acid is acetic acid, butyric acid, or a mixture of both. The fermentation method provides a simultaneous method of producing high specific productivity of oxygen-containing hydrocarbon compounds and producing nutrient supplements from the bacterial cells used in the fermentation method. Herein, specific productivity is expressed as specific STY. In this embodiment, specific oxygen-containing hydrocarbon compound productivity can be expressed as specific STY (for example, specific air-hour yield can be expressed as alcohol (g) / day / cell 1 gram or organic acid (g) / day / cell 1 gram). In one embodiment, the fermentation method yields a specific organic acid production of approximately 0.2 to 50 grams of organic acid per day per gram of cell; in another embodiment, approximately 0.2 to 20 grams of organic acid per day per gram of cell; in yet another embodiment, approximately 10 to 50 grams of organic acid per day per gram of cell; in yet another embodiment, approximately 14 to 30 grams of organic acid per day per gram of cell; in yet another embodiment, approximately 2 to 20 grams of organic acid per day per gram of cell; and in yet another embodiment, approximately 15 to 25 grams of organic acid per day per gram of cell. In this embodiment, the organic acid is acetic acid, butyric acid, or a mixture of both.

[0046] The organic acid 227 produced from the acetic acid-producing CO2 fermentation 220 can be sent to the CO fermentation 230 to increase the alcohol production capacity in the CO fermentation 230. In this embodiment, CO fermentation yields a specific alcohol production capacity of approximately 16 grams or more of alcohol per day per cell, in another embodiment, a specific alcohol production rate of approximately 18 g or more per day per cell, in another embodiment, a specific alcohol production rate of approximately 20 g or more per day per cell, in another embodiment, a specific alcohol production rate of approximately 22 g or more per day per cell, in another embodiment, a specific alcohol production rate of approximately 24 g or more per day per cell, in another embodiment, a specific alcohol production rate of approximately 16 to approximately 30 g per day per cell, in another embodiment, approximately 18 to approximately 34 g per day per cell, in another embodiment, approximately 20 to approximately 40 g per day per cell, in another embodiment, approximately 22 to approximately 48 g per day per cell, in another embodiment, approximately 20 to approximately 50 g per day per cell, in another embodiment, approximately 25 to approximately 50 g per day per cell, and in another embodiment, approximately 25 to approximately 55 g per day per cell. In this embodiment, the alcohol is ethanol, butanol, or a mixture of both.

[0047] The acetic acid production CO2 fermentation method can yield CO2 conversion rates of 75% or more in one embodiment, 80% or more in one embodiment, 85% or more in one embodiment, 90% or more in one embodiment, 85% to 95% in another embodiment, and 90% to 99% in yet another embodiment. When the acetic acid production CO2 fermentation vessel 220 is connected to the CO fermentation vessel 230 and the methane-producing bacteria fermentation vessel 105, the CO2 conversion rate in the acetic acid production CO2 fermentation vessel 220 may be intentionally reduced in order to maintain a balance between organic acid consumption in the CO fermentation vessel 230 and CO2 consumption in the acetic acid production CO2 fermentation vessel 220 and the methane-producing bacteria fermentation vessel 105. In this scenario, the CO2 conversion rate of acetic acid production CO2 fermentation is controlled to 25% to 74%, 45% to 74% in one embodiment, 55% to 75% in one embodiment, 60% to 74% in one embodiment, 62% to 74% in one embodiment, and 65% to 74% in another embodiment. By controlling the CO2 conversion rate, the vent gas 225 produced from the acetic acid production CO2 fermentation vessel 220 can contain 15-45% CO2. The vent gas 225 is then supplied to the methane-producing bacteria fermentation vessel 105 to produce methane.

[0048] Alternatively, to maintain a balance in organic acid consumption in the CO fermentation vessel 230, vent gas 481 can be supplied from the CO fermentation vessel 230 to both the acetic acid-producing CO2 fermentation vessel 220 and the methane-producing bacteria fermentation vessel 105. In this scenario, the CO2 conversion rate in the acetic acid-producing CO2 fermentation vessel 220 is maintained at 75-99%. The vent gas 225 produced from the high-CO2-conversion-rate acetic acid-producing CO2 fermentation vessel 220 has a CO2 concentration of 2-15%. Optionally, the vent gas 225 can be supplied to the methane-producing bacteria fermentation vessel 105 to produce methane. Furthermore, the fermentation method may be operated under conditions that promote the production of the desired product. In one embodiment, the desired product is one or more oxygen-containing hydrocarbon compounds. In another embodiment, the desired product is the microbial biomass itself, and the method produces the oxygen-containing hydrocarbon compounds as byproducts. Operating parameters such as culture medium flow rate, gaseous substrate feed rate, water supply / recycle rate, temperature, medium oxidation-reduction potential, pressure, pH, stirring rate (if using a stirred-tank reactor), and cell concentration are monitored and controlled throughout the fermentation method.

[0049] When the acetic acid production CO2 fermentation method begins, a fermented liquid broth is produced inside the fermentation vessel. In addition to the culture medium, the fermented liquid broth also contains acetic acid-producing bacteria and one or more oxygen-containing hydrocarbon compounds. Generally, the fermented liquid broth has a pH of about 8 or less, or in another embodiment, about 7.5 or less. In one embodiment, the cell concentration of the fermented liquid broth is about 1 to about 15 g / L, in another embodiment, 2 to about 30 g / L, in another embodiment, about 2 to about 25 g / L, in another embodiment, about 2 to about 20 g / L, in another embodiment, about 2 to about 10 g / L, in another embodiment, about 2 to about 8 g / L, in another embodiment, about 3 to about 30 g / L, in another embodiment, about 3 to about 9 g / L, and in another embodiment, about 4 to about 8 g / L.

[0050] CO fermentation As described herein in relation to Figure 2, certain acetic acid-producing bacteria can ferment a CO-containing gaseous substrate 50 into useful oxygen-containing hydrocarbon compounds such as ethanol and butanol 255, and Clostridium fermentation liquid broth 270. Acetic acid-producing bacteria in the CO fermentation vessel 230 can also convert organic acids into oxygen-containing hydrocarbon compounds in the presence of CO. As shown in Figure 3, the vent gas 481 from the CO fermentation vessel 230 can be returned to the acetic acid-producing CO2 fermentation vessel 220 and / or the methane-producing bacteria fermentation vessel. In this embodiment, the vent gas 481 from the CO fermentation vessel 230 can be processed to remove CO before entering the methane-producing bacteria fermentation vessel 105. Integrated fermentation system As those skilled in the art will understand, one or all of the methods described in Figures 1-3 can be combined into the overall system. Microbial biomass The fermentation liquid broth from any of the fermentation vessels (Figure 1: 140, Figure 2: 140 and 270, Figure 3: 140, 250 and 270) can be further purged from the fermentation vessel and then separated into a cell-free permeate and a cell-containing suspension using one or more cell separation devices. The cell membranes of the cell-containing suspension are ruptured to generate homogenates. Using a fractionation device, the homogenates are fractionated into a protein-containing supernatant and a protein-containing cell debris portion.

[0051] Suitable cell separation devices include, but are not limited to, filtration devices, hollow fiber filtration devices, spiral filtration devices, ultrafiltration devices, ceramic filter devices, cross-flow filtration devices, size exclusion column filtration devices, spiral membranes, centrifugal devices, and combinations thereof. A method for producing single-cell proteins from biomass is described in U.S. Patent Application No. 16 / 416,133, filed May 17, 2019. The cell-containing suspension contains microbial cells at a higher cell concentration than the fermented liquid broth. In one embodiment, the cell concentration of the cell-containing suspension is approximately 20 g or more / L, in another embodiment, approximately 30 g or more / L, in another embodiment, approximately 40 g or more / L, in another embodiment, approximately 50 g or more / L, in another embodiment, approximately 60 g or more / L, in another embodiment, approximately 20 to approximately 300 g / L, in another embodiment, approximately 30 to approximately 250 g / L, in another embodiment, approximately 40 to approximately 200 g / L, in another embodiment, approximately 50 to approximately 150 g / L, and in yet another embodiment, approximately 100 to approximately 150 g / L.

[0052] The cells in the cell-containing suspension may be ruptured using one or more rupture devices selected from the group consisting of microfluidic devices, sonic devices, ultrasonic devices, mechanical rupture devices, French presses, freezers, heaters, heat exchangers, distillation columns, sterilization devices, UV sterilization devices, gamma ray sterilization devices, reactors, homogenizers, and combinations thereof. In one embodiment, the pH of the cell-containing suspension is adjusted to about 6 to about 12 before rupturing the cell membranes of the cell-containing suspension; in another embodiment, 7 to 12; in another embodiment, 8 to 12; in another embodiment, 7.5 to 11; in yet another embodiment, 8.5 to 11; and in yet another embodiment, 7 to 10.

[0053] In another embodiment, the cell-containing suspension is hydrolyzed by a hydrolase enzyme. In this embodiment, the cell-containing suspension and the hydrolase enzyme are incubated at a temperature of about 50 to about 70°C for about 3 to about 72 hours, in one embodiment 3 to 48 hours, in one embodiment 4 to 24 hours, in one embodiment 6 to 24 hours, in another embodiment 6 to 12 hours, and in yet another embodiment 4 to 12 hours to form a hydrolyzed lysate. The pH of the cell-containing suspension is adjusted to about 6 to about 12 pH, in another embodiment 7 to 12 pH, in another embodiment 8 to 12 pH, in another embodiment 7.5 to 11 pH, in another embodiment 8.5 to 11 pH, and in yet another embodiment 7 to 10 pH before the hydrolysis of the cell-containing suspension. The hydrolase enzyme is selected from the group consisting of subyrase, alcalase, serine protease, serine endopeptidase, and mixtures thereof. The hydrolyzed lysate is fractionated into a protein-containing supernatant and a protein-containing cell debris portion using centrifugation, ultrafiltration, and / or a combination thereof. The protein-containing supernatant has a nucleic acid content of less than approximately 5%, less than 4% in one embodiment, less than 3% in another embodiment, and less than 2% in another embodiment.

[0054] The protein-containing supernatant and protein-containing cell debris can be used as is or further processed into protein-containing nutritional supplements. A dehydration unit can be used to dry the protein-containing supernatant and produce soluble protein-containing nutritional supplements such as protein powder. Suitable dehydration units include spray drying units, drum drying units, freeze-drying units, lyophilizing units, and combinations thereof. The protein-containing supplement can be purified by further removal of other components such as water and ash. The protein-containing supplement can be used as animal feed as is, or it can be blended with other materials to produce one or more types of nutritional supplements. In one embodiment, the protein-containing supplement contains about 60 to about 99 mass percent protein; in another embodiment, about 70 to about 95 mass percent protein; in another embodiment, about 75 to about 95 mass percent protein; in another embodiment, about 80 to about 95 mass percent protein; and in another embodiment, about 85 to about 95 mass percent protein. [Examples]

[0055] The following embodiments further illustrate the present disclosure and should not be construed as limiting its scope. (Example 1) Methanothermobacter thermautotrophicus fermentation CO2 and H2 gases are continuously introduced into a stirred tank bioreactor containing Metanothermobacter thermautotrophus along with a standard liquid culture medium containing trace metals and salts. Vitamins are supplied using a dedicated feed line.

[0056] The New Brunswick Bioflow 320 reactor containing the fermentation medium is started with actively growing Metanothermobacter thermautotrophus. The reactor stirring speed is set to 1200 rpm at the start of the experiment. This stirring speed is maintained throughout the experiment. The feed gas flow to the reactor is increased based on the absorption of H2 and CO2 by the culture. The bioreactor temperature is maintained at approximately 60°C throughout the experiment. Samples of the gas feed to the bioreactor, the off-gas from the bioreactor, and the fermentation broth in the bioreactor are taken at intervals. For example, feed gas, off-gas, and fermentation broth are sampled approximately daily, every 2 hours, and every 4 hours, respectively. The above samples are analyzed for the consumption or production of various gas components and the optical density (cell density) of the culture. The stationary volume of the reactor, i.e., the volume of fermentation broth, is maintained at approximately 2000–2200 ml throughout the experiment. Furthermore, a mass flow controller is used to regulate the gas flow to the reactor, allowing for real-time measurement of the gas flow. The feed gas composition for this experiment is 76% H2, 20% CO2, and 4% N2.

[0057] In this experiment, a cell recycling system (CRS) is attached to the reactor before starting the experiment. During the experiment, the flow rate of nutrients (growth medium) to the reactor is 2.0–5.0 ml / min. The culture purging rate is 3.0–5.0 ml / min, and the permeate is removed through the CRS at a rate of 0–2.0 ml / min. The results can be summarized as follows. Specific CO2 absorption: 0.8-1.1 mmol CO2 / min / gram of dry cells Ratio H2 absorption: 3.3-3.8 mmol H2 / min / gram of dry cells Average cell retention time: 11.2 hours Average cell density: 3g / L CO2 conversion rate: 90%~99% The efflutent gas composition in this experiment is 13.8% H2, 7.6% CO2, 62.5% CH4, and 16.1% N2. The specific methane production capacity is 0.77 mmol / min / cell / gram.

[0058] (Example 2) Methanothermobacter marburgensis fermentation CO2 and H2 gases are continuously introduced into a stirred tank bioreactor containing Metanothermobacter marburgensis along with a standard liquid culture medium containing trace metals and salts. Vitamins are supplied using a dedicated feed line.

[0059] The New Brunswick Bioflow 310 reactor containing the fermentation medium is started with actively growing Metanothermobacter marbulgensis. The reactor stirring speed is set to 1200 rpm at the start of the experiment. This stirring speed is maintained throughout the experiment. The feed gas flow to the reactor is increased based on the absorption of H2 and CO2 by the culture. The bioreactor temperature is maintained at approximately 60°C throughout the experiment. Samples of the gas feed to the bioreactor, the off-gas from the bioreactor, and the fermentation broth in the bioreactor are taken at intervals. For example, feed gas, off-gas, and fermentation broth are sampled approximately daily, every 2 hours, and every 4 hours, respectively. The above samples are analyzed for the consumption or production of various gas components and the optical density (cell density) of the culture. The stationary volume of the reactor is maintained at approximately 1500-1600 ml throughout the experiment. The gas flow to the reactor is also measured in real time using a mass flow controller that regulates the gas to the reactor. The feed gas composition for this experiment is 70% H2, 25% CO2, and 5% N2.

[0060] During the experiment, the flow rate of nutrients (growth medium) into the reactor is 1.0-2.0 ml / min, and 1.0-2.0 ml / min of culture is removed from the reactor through a culture purge pump. The results can be summarized as follows. Specific CO2 absorption: 0.6-0.9 mmol CO2 / min / gram of dry cells Ratio H2 absorption: 2.8-3.2 mmol H2 / min / gram of dry cells Average cell retention time: 18.2 hours Average cell density: 3.4g / L CO2 conversion rate: 65%~85% The efferent gas composition for this experiment was 7.4% H2, 26.8% CO2, 49.1% CH4, and 16.7% N2. The specific methane production capacity is 0.67 mmol / min / cell / gram.

[0061] (Example 3) Effluent gas reforming A portion of the efferent gas produced in Example 1 is sent to a methane reformer that converts methane into CO and H2. CH4+H2O(+heat)→CO+3H2 The reformed gas composition is 21.7% CO, 69.8% H2, 2.7% CO2, 0.2% CH4, and 5.6% N2. Next, an H2 removal unit separates the reformed gas into a CO-rich synthesis gas having 68.6% CO, 4.4% H2, 8.4% CO2, 0.7% CH4, and 17.9% N2, and an H2-rich stream having a 99% H2 concentration.

[0062] (Example 4) Effient gas pyrolysis A portion of the efferent gas produced in Example 1 is sent to a gas separation unit to produce a methane-enriched feed gas having 2.6% H2, 0.7% CO2, 93.7% CH4, and 3% N2, and a methane-diluted synthesis gas having 52% H2, 32.6% CO2, 6.7% CH4, and 8.7% N2. The methane-enriched feed gas is then sent to a microwave pyrolysis cracking unit to produce H2 and elemental carbon. CH4(+heat)→C+2H2 (Example 5) Clostridium jungdarii and Methanothermobacter sarmuottrophus fermentation A synthesis gas containing CO, CO2, and H2 is continuously introduced into a stirred tank bioreactor containing Clostridium lyngdary along with a liquid culture medium containing trace metals and salts as described herein. Vitamins are supplied using a dedicated feed line.

[0063] The New Brunswick Bioflow reactor containing the fermentation medium is started with actively growing Clostridium lyngdaryi. The reactor stirring speed is set to 800 rpm at the start of the experiment and this stirring speed is maintained throughout the experiment. The feed gas flow to the reactor is increased based on the absorption of H2 and CO from the culture. The bioreactor temperature is maintained at approximately 38°C throughout the experiment. Samples of the gas feed to the bioreactor, the off-gas from the bioreactor, and the fermentation broth in the bioreactor are taken at intervals. For example, feed gas, off-gas, and fermentation broth are sampled approximately daily, every 2 hours, and every 4 hours, respectively. The above samples are analyzed for the consumption or production of various gas components, broth acetic acid concentration, broth ethanol concentration, and the optical density (cell density) of the culture. The stationary volume of the reactor is maintained at 3000–3250 ml throughout the experiment. Furthermore, a mass flow controller is used to maintain the gas flow to the reactor at the required average gas flow rate of 67.0 mmol / min. The feed synthesis gas composition is 23% H2, 35% CO, 29% CO2, and 13% N2.

[0064] The following results are achieved during fermentation. Specific CO absorption: 0.9-1.3 mmol CO / min / cell per gram Relative H2 absorption: 0.2-0.6 mmol H2 / min / cell per gram Average cell density: 5.85g / L Average cell retention time: 8 hours CO conversion rate: 90% H2 conversion rate: 30% Specific ethanol production capacity: 13.31 grams / day / 1 gram per cell Vent gas composition: 4% CO, 20% H2, 59% CO2, and 16% N2 Vent gas flow rate: 52.95 mmol / min

[0065] Next, the vent gas is blended with the H2 stream and fed into a methanothermobacter thermautotrophus fermentation reactor. The methanothermobacter thermautotrophus fermentation is carried out as illustrated in Example 1. The two-reactor method is configured as shown in Figure 2. Blended feed gas composition: 1.4% CO, 74.9% H2, 18.5% CO2, and 5.2% N2.

[0066] The following results are achieved during fermentation. Specific CO2 absorption: 0.8-1.3 mmol CO2 / min / cell per gram Ratio H2 absorption: 3.2-4.5 mmol H2 / min / cell per gram Average cell retention time: 10 hours Average cell density: 3g / L CO2 conversion rate: 90%~99% Specific methane production rate: 0.83 mmol / min / cell / gram Effient gas composition: 9.8% H2, 1.7% CO2, 67.6% CH4, and 20.9% N2. Effient gas flow rate: 7.4 mmol / min The average gas composition and gas flow rate are as follows: [Table 4]

[0067] Next, a portion of the efferent gas can be sent to a methane reformer as illustrated in Example 3 to produce CO-rich synthesis gas and H2-rich flow. The CO-rich synthesis gas can then be blended with synthesis gas to supply the blended synthesis gas feed gas to a Clostridium lyngdary fermentation reactor, and the H2-rich flow can be used to blend with vent gas from Clostridium lyngdary fermentation. The following points are illustrated by a comparison of percentages in gas composition: The percentage of CO ranging from a high of 35% to a low of 0%. In this embodiment, the CO reduction percentage can range from 90-100%, in another embodiment from 95-100%, and in yet another embodiment from 99-100%. The percentage of CO2 is from a high of 29% to a low of 1.7%. In this embodiment, the percentage reduction in CO2 can be 90-95%, in another embodiment, 92-95%, and in yet another embodiment, 94-95%.

[0068] (Example 6) Fermentation of Clostridium lyngdaryi, Acetobacterium woodyi, and Methanothermobacter sarmototropicus A New Brunswick Bioflow bioreactor, containing Acetobacterium woodie actively growing with the growth medium, is added to the system exemplified in Example 5 to receive vent gas from the Clostridium lyngdaryi fermentation bioreactor. The stirring speed of the Acetobacterium woodie bioreactor is set to 600 rpm. This stirring speed is kept constant throughout the experiment. The feed gas flow to the reactor is maintained at 36.6 ml / min to 44.4 ml / min. The bioreactor temperature is maintained at approximately 33°C throughout the experiment. + Maintain a gas level of 3500–4000 ppm. Samples of gas feed to the bioreactor, off-gas from the bioreactor, and fermentation broth in the bioreactor are taken at intervals. For example, feed gas, off-gas, and fermentation broth are sampled approximately daily, every 2 hours, and every 4 hours, respectively. The above samples are analyzed for the consumption or production of various gas components, broth acetate concentration, and optical density (cell density) of the culture. The stationary volume of the reactor was maintained at 1900–2275 ml throughout the experiment. Furthermore, the gas flow to the reactor was maintained at the required gas flow rate by using a mass flow controller.

[0069] Before starting the experiment, attach the Cell Recycling System (CRS) to the reactor. During the experiment, maintain a nutrient (growth medium) flow rate of 2.8 ml / min to the reactor. Maintain the medium feed rate throughout the experiment. The average rate requirement for base (NaOH) to maintain pH 6.5 is 0.075 ml / min, and remove 2.9 ml / min of permeate from the reactor through the CRS.

[0070] Vent gas from the Acetobacterium woodii bioreactor is blended with a supplement H2 stream and sent to the Metanothermobacter thermautotrophus fermentation bioreactor. Acetic acid produced from the Acetobacterium woodii bioreactor is sent to the Clostridium ljundahlii fermentation bioreactor. The CO2 conversion rate of the Acetobacterium woodii fermentation initially reached 90-97%. After closing the water loop between the Acetobacterium woodii bioreactor and the Clostridium ljundahlii bioreactor, the CO2 conversion rate of the Acetobacterium woodii fermentation was intentionally reduced to 60-70%. The Metanothermobacter thermautotrophus fermentation and Clostridium ljundahlii fermentation are carried out as illustrated in Example 5. The three-reactor method is configured as shown in Figure 3. Feed gas composition for Clostridium lyngdary fermentation: 36% CO, 63% H2, 0% CO2, and 1% N2

[0071] The following results are achieved during Clostridium lyngdarie fermentation. Specific CO absorption: 1.3-1.7 mmol CO / min / cell per gram Relative H2 absorption: 0.2-0.6 mmol H2 / min / cell per gram Average cell density: 6.43g / L Average cell retention time: 7.2 hours CO conversion rate: 90% H2 conversion rate: 14% Specific ethanol production capacity (before water loop closure): 13.43 grams / day / 1 gram per cell Specific ethanol production capacity (after water loop closure): 36.87 grams / day / 1 gram per cell Vent gas composition: 4% CO, 67% H2, 27% CO2, and 1% N2 Vent gas flow rate: 71.77 mmol / min

[0072] The following results are achieved during Acetobacterium woodyi fermentation. Specific CO2 absorption: 0.4-0.9 mmol CO / min / cell per gram Ratio H2 absorption: 1.0-1.5 mmol H2 / min / cell per gram Average cell density: 6.3g / L Average cell retention time: 20 hours CO conversion rate: 100% CO2 conversion rate: 60% H2 conversion rate: 48% Specific acetic acid production capacity: 30.55 grams / day / 1 gram per cell Vent gas composition: 0% CO, 74.6% H2, 22.8% CO2, and 2.6% N2 Vent gas flow rate: 33.84 mmol / min

[0073] The following results are achieved during the fermentation of Methanothermobacter thermautotrophus. Specific CO2 absorption: 0.6-1.1 mmol CO / min / cell per gram Ratio H2 absorption: 3.3-4.0 mmol H2 / min / cell per gram Average cell density: 4.24g / L Average cell retention time: 11 hours CO2 conversion rate: 99% H2 conversion rate: 99% Specific methane production rate: 0.77 mmol / min / cell / gram Effient gas composition: 0% CO, 4% H2, 1% CO2, 84% CH4, and 11% N2 Effluent gas flow rate: 7.86 mmol / min

[0074] The average gas composition and gas flow rate are as follows: [Table 5]

[0075] The following points are illustrated by a comparison of percentages in gas composition: The percentage of CO ranging from a high of 36% to a low of 0%. In this embodiment, the CO reduction percentage can range from 90-100%, in another embodiment from 95-100%, and in yet another embodiment from 99-100%. The percentage of CO2 from a high of 27% to a low of 1%. In this embodiment, the percentage reduction in CO2 can be 90-97%, in another embodiment, 95-97%, and in yet another embodiment, 96-97%.

[0076] While the disclosure disclosed herein has been described by specific embodiments, examples, and applications thereof, other modifications and variations can be made without departing from the basic scope of the disclosure as defined in the claims.

Claims

1. CO 2 A method of converting, CO 2 and H 2 A step of fermenting a gaseous substrate containing in a methane-producing microbial fermentation vessel with methane-producing archaea, wherein the gaseous substrate contains CO2 in a ratio of approximately 1:3 to approximately 1:

4. 2 vs H 2 The ratio of steps is maintained, The steps include recovering methane from the methane-producing microbial fermentation vessel and A method that includes this.

2. Methanogenic archaea include Methanobacterium alcaliphilum, Methanobacterium bryantii, Methanobacterium congolense, Methanobacterium defluvii, Methanobacterium espanolae, Methanobacterium formicicum, Methanobacterium ivanovii, Methanobacterium palustre, Methanobacterium thermaggregans, and Methanobacterium euriginosum. Methanobrevibacter uliginosum, Methanobrevibacter acididurans, Methanobrevibacter arboriphilicus, Methanobrevibacter gottschalkii, Methanobrevibacter olleyae, Methanobrevibacter ruminantium, Methanobrevibacter smithii, Methanobrevibacter woesei, Methanobrevibacter wolinii, Methanothermobacter Methanothermobacter marburgensis, Methanothermobacter thermautotrophumMethanothermobacter thermoflexus, Methanothermobacter thermophilus, Methanothermobacter wolfeii, Methanothermus sociabilis, Methanocorpusculum bavaricum, Methanocorpusculum parvum, Methanoculleus chikuoensis, Methanoculleus submarinus, Methanogenium frigidum, Methanogenium liminatans, Methanogenium marinum Methanosarcina marinum, Methanosarcina acetivorans, Methanosarcina barkeri, Methanosarcina mazei, Methanosarcina thermophila, Methanomicrobium mobile, Methanocaldococcus jannaschii, Methanococcus aeolicus, Methanococcus maripaludis, Methanococcus vannielii, Methanococcus voltaei, MethanothermococcusThe method according to claim 1, selected from the group consisting of *Methanocaldococcus thermolithotrophicus*, *Methanopyrus kandleri*, *Methanothermobacter thermoautotroiphicus*, *Methanocaldococcus fervens*, *Methanocaldococcus indicus*, *Methanocaldococcus infernus*, and *Methanocaldococcus vulcanius*.

3. The method according to claim 1, wherein the methanogenic archaeon is metanothermobacter thermoautotrophycus.

4. Approximately 0.5 to 3 mmol of CO 2 / min / cell ratio CO2 2 The method according to claim 1, which provides absorption.

5. A ratio of H of about 1.5 to about 12 mmol 2 / minute / gram of cells 1 that results in H 2 absorption, the method according to claim 1.

6. The method according to claim 1, having a cell retention time of approximately 5 to approximately 50 hours.

7. The method according to claim 1, which provides a methane production capacity of approximately 0.5 to approximately 2.5 mmol of methane / min / cell and 1 gram of methane.

8. More than 65% CO 2 The method according to claim 1, which provides a conversion rate.

9. The steps include obtaining a fermentation liquid broth containing methanogenic archaea from a methanogenic microbial fermentation vessel, The steps include separating the fermented liquid broth into a cell-free permeate and a cell-containing suspension, The steps include: rupturing the cell membrane of a cell-containing suspension to generate a homogenate; The steps include: using a fractionation apparatus to fractionate the homogenate into a protein-containing supernatant and a protein-containing cell debris portion; Steps to obtain protein-containing nutritional supplements and The method according to claim 1, further comprising:

10. The method according to claim 9, wherein the cell-containing suspension has a dry cell mass concentration of about 50 g / liter to about 200 g / liter.

11. The method according to claim 9, wherein the step of rupturing the cell membrane of a cell-containing suspension is performed using one or more rupture devices selected from the group consisting of microfluidic devices, sonic processing devices, ultrasonic devices, mechanical destruction devices, French presses, freezers, heaters, heat exchangers, distillation columns, sterilization devices, UV sterilization devices, gamma ray sterilization devices, reactors, homogenizers, and combinations thereof.

12. The method according to claim 9, wherein, prior to the step of rupturing the cell membranes of the cell-containing suspension, the pH of the cell-containing suspension is adjusted to approximately 6 to approximately 12.

13. The method according to claim 9, wherein the homogenate is a hydrolyzed lysate formed by contacting a cell-containing suspension with a hydrolase enzyme.

14. The method according to claim 13, wherein a cell-containing suspension and a hydrolase enzyme are incubated at a temperature of about 50 to about 70°C for about 3 to about 72 hours to form a hydrolyzed lysate.

15. The method according to claim 13, wherein the hydrolase enzyme is selected from the group consisting of subyrase, alcalase, serine protease, serine endopeptidase, and mixtures thereof.

16. The method according to claim 9, wherein the homogenate is fractionated into a protein-containing supernatant and a protein-containing cell debris portion using centrifugation, ultrafiltration, and / or a combination thereof.

17. The method according to claim 9, wherein the protein-containing supernatant has a nucleic acid content of less than approximately 5%.

18. The method according to claim 9, wherein the protein-containing supernatant is dehydrated to obtain a soluble protein-containing nutritional supplement containing about 60 to about 99% dry mass of protein.

19. CO 2 A method of converting, CO 2 and H 2 The steps include fermenting a gaseous substrate containing methane-producing archaea in a methane-producing fermentation vessel, The steps include recovering methane from the methane-producing microbial fermentation vessel, By cracking at least a portion of the methane, H 2 The steps to generate, H 2 The steps include returning at least a portion of it to the methane-producing bacteria fermentation vessel and A method that includes this.

20. In a gaseous substrate, CO is present in a ratio of approximately 1:3 to approximately 1:

4. 2 vs H 2 The method according to claim 19, wherein the ratio is maintained.

21. The method according to claim 19, wherein the methanogenic archaeon is metanothermobacter thermoautotrophycus.

22. Approximately 0.5 to 3 mmol of CO 2 / min / cell ratio CO2 2 The method according to claim 19, which provides absorption.

23. Approximately 1.5 to 12 mmol of H 2 / min / cell-to-gram ratio H 2 The method according to claim 19, which provides absorption.

24. The method according to claim 19, having a cell retention time of approximately 5 to approximately 50 hours.

25. The method according to claim 19, which provides a methane production capacity of approximately 0.5 to approximately 2.5 mmol of methane / min / cell and 1 gram of methane.

26. More than 65% CO 2 The method according to claim 19, which provides a conversion rate.

27. The method according to claim 19, wherein the cracking is performed using a methane cracking device selected from the group consisting of a microwave pyrolysis cracking device, a molten metal pyrolysis cracking device, a plasma arc pyrolysis cracking device, and a combination thereof.

28. The method according to claim 19, wherein the cracking is carried out using a methane cracking apparatus selected from the group consisting of a steam reformer, a dry reformer, a partial oxidation reformer, and a combination thereof.

29. The steps include obtaining a fermentation liquid broth containing methanogenic archaea from a methanogenic microbial fermentation vessel, The steps include separating the fermented liquid broth into a cell-free permeate and a cell-containing suspension, The steps include: rupturing the cell membrane of a cell-containing suspension to generate a homogenate; The steps include: using a fractionation apparatus to fractionate the homogenate into a protein-containing portion and a protein-containing cell debris portion; Steps to obtain protein-containing nutritional supplements and The method according to claim 19, further comprising:

30. CO and CO 2 A method of converting, CO and H 2 A gaseous substrate containing CO is fermented in a CO fermentation vessel with CO-converting acetic acid-producing bacteria to produce alcohol and CO 2 The steps include producing the contained vent gas, CO 2 The steps include providing the contained vent gas from the CO fermentation vessel to the methane-producing bacteria fermentation vessel, CO 2 The process involves fermenting the contained vent gas with methane-producing archaea in a methane-producing microbial fermentation vessel to produce methane. A method that includes this.

31. The method according to claim 30, wherein the gaseous substrate contains at least 20 mol% CO.

32. Methanogenic archaea include Metanobacterium alkaliphyllum, Metanobacterium bryantyi, Metanobacterium congolense, Metanobacterium deflubyi, Metanobacterium espanolae, Metanobacterium formicicum, Metanobacterium ibanobii, Metanobacterium parstre, Metanobacterium thermagregans, Metanobacterium euriginosum, Metanobrevibacter acididurans, and Metanobrevibacter - Arboriphyllicus, Metanobrevibacter gotscharkii, Metanobrevibacter oreyae, Metanobrevibacter luminantium, Metanobrevibacter smisi, Metanobrevibacter oozei, Metanobrevibacter wolinii, Metanothermobacter marbulgensis, Metanothermobacter thermoautotrophum, Metanothermobacter thermoflexus, Metanothermobacter thermophilus, Metanothermobacter wolfei Metanotermus sochiabilis, Metanocorpus bavaricum, Metanocorpus palham, Metanocreus thikoensis, Metanocreus submarinus, Metanogenium frigidum, Metanogenium liminatans, Metanogenium marinum, Metanosarcinia acetylborans, Metanosarcinia barkerii, Metanosarcinia mazei, Metanosarcinia thermophylla, Metanomicrobium mobile, Metanocardococcus janaskii, Metanoco The method according to claim 30, selected from the group consisting of Cass aeolicus, Metanococcus maripaldis, Metanococcus van niirii, Metanococcus voltae, Metanothermococcus thermolotrophus, Metanopyrus candrelli, Metanothermobacter thermoautotrophycus, Metanocardococcus favens, Metanocardococcus indicus, Metanocardococcus infernus, and Metanocardococcus bulcanius.

33. The method according to claim 30, wherein the methanogenic archaeon is metanothermobacter thermoautotrophycus.

34. The methane-producing microbial fermentation vessel contains approximately 1 to 3 mmol of CO2. 2 / min / cell ratio CO2 2 The method according to claim 30, which provides absorption.

35. The methane-producing bacteria fermentation vessel contains approximately 3 to 12 mmol of H. 2 / min / cell-to-gram ratio H 2 The method according to claim 30, which provides absorption.

36. The method according to claim 30, wherein the methane-producing microbial fermentation vessel has a cell retention time of about 5 to about 50 hours.

37. The method according to claim 30, wherein the methane-producing microbial fermentation vessel yields a methane production capacity of approximately 1.2 to approximately 2.5 mmol of methane / minute / cell per gram.

38. The methane-producing bacteria fermentation vessel contains more than 65% CO2. 2 The method according to claim 30, which provides a conversion rate.

39. CO-converting acetic acid-producing bacteria include Clostridium aceticum, Clostridium acetobutylicum, Clostridium acetobutylicum P262, Clostridium autoethanogenum (DSM 19630, German DSMZ), Clostridium autoethanogenum (DSM 10061, German DSMZ), Clostridium autoethanogenum (DSM 23693, German DSMZ), Clostridium autoethanogenum (DSM 24138, German DSMZ), and Clostridium carboxydivorans. carboxidivorans), Clostridium coskatii (ATCC PTA-10522), Clostridium drakei, Clostridium ljungdahlii PETC (ATCC 49587), Clostridium ljungdahlii ERI2 (ATCC 55380), Clostridium ljungdahlii C-01 (ATCC 55988), Clostridium ljungdahlii O-52 (ATCC 55889), Clostridium magnum, Clostridium pasteurianum (DSM) 525, German DSMZ), Clostridium ragsdalei P11 (ATCC BAA-622), Clostridium scatogenesThe method according to claim 30, selected from the group consisting of Clostridium scatologenes, Clostridium thermoaceticum, Clostridium ultunense, Clostridium stick-landii, and mixtures thereof.

40. The method according to claim 30, wherein the alcohol is ethanol.

41. The method according to claim 30, wherein the CO fermentation vessel yields a CO conversion rate of 80% or more.

42. Methane is supplied from the methane-producing bacteria fermentation vessel to the methane cracking device, H 2 Steps include producing rich flow and CO-rich synthesis gas, The steps include supplying CO-rich synthesis gas to a CO fermentation vessel, H 2 The step of supplying the rich flow to the methane-producing bacteria fermentation vessel and The method according to claim 30, further comprising:

43. Methane is supplied from the methane-producing bacteria fermentation vessel to the methane cracking device, H 2 and a step of producing a CO-containing gaseous substrate, H 2 and the step of supplying a CO-containing gaseous substrate to a CO fermentation vessel. The method according to claim 30, further comprising:

44. The steps include obtaining a fermentation liquid broth containing CO-converting acetic acid-producing bacteria from a CO-fermentation vessel, and obtaining a fermentation liquid broth containing methane-producing archaea from a methane-producing bacteria fermentation vessel, The process involves separating a fermented liquid broth containing CO-converting acetic acid-producing bacteria and a fermented liquid broth containing methane-producing archaea into a cell-free permeate and a cell-containing suspension. The steps include: rupturing the cell membrane of a cell-containing suspension to generate a homogenate; The steps include: using a fractionation apparatus to fractionate the homogenate into a protein-containing supernatant and a protein-containing cell debris portion; Steps to obtain protein-containing nutritional supplements and The method according to claim 30, further comprising:

45. The method according to claim 44, wherein the cell-containing suspension has a dry cell mass concentration of about 20 g / liter to about 200 g / liter.

46. The method according to claim 44, wherein the step of rupturing the cell membrane of a cell-containing suspension is performed using one or more rupture devices selected from the group consisting of microfluidic devices, sonic processing devices, ultrasonic devices, mechanical destruction devices, French presses, freezers, heaters, heat exchangers, distillation columns, sterilization devices, UV sterilization devices, gamma ray sterilization devices, reactors, homogenizers, and combinations thereof.

47. The method according to claim 44, wherein, before the step of rupturing the cell membranes of the cell-containing suspension, the pH of the cell-containing suspension is adjusted to about 6 to about 12.

48. The method according to claim 44, wherein the homogenate is a hydrolyzed lysate formed by contacting a cell-containing suspension with a hydrolase enzyme.

49. The method according to claim 48, wherein a cell-containing suspension and a hydrolase enzyme are incubated at a temperature of about 50 to about 70°C for about 3 to about 72 hours to form a hydrolyzed lysate.

50. The method according to claim 48, wherein the hydrolase enzyme is selected from the group consisting of subyrase, alcalase, serine protease, serine endopeptidase, and mixtures thereof.

51. The method according to claim 44, wherein the homogenate is fractionated into a protein-containing supernatant and a protein-containing cell debris portion using centrifugation, ultrafiltration, and / or a combination thereof.

52. The method according to claim 44, wherein the protein-containing supernatant has a nucleic acid content of less than about 5%.

53. The method according to claim 44, wherein the protein-containing supernatant is dehydrated to obtain a soluble protein-containing nutritional supplement containing about 60 to about 99% dry mass of protein.

54. CO and CO 2 A method of converting, CO and H 2 A gaseous substrate containing CO is fermented in a CO fermentation vessel with CO-converting acetic acid-producing bacteria to produce alcohol and the first CO 2 The steps include producing the contained vent gas, First CO 2 The contained vent gas is converted into acetic acid CO from the CO fermentation vessel. 2 Steps for providing to the fermentation vessel, First CO 2 The contained vent gas is converted into acetic acid CO2. 2 CO2 in the fermentation vessel 2 Fermented with acetic acid-producing bacteria, organic acids and a second CO2 are produced. 2 The steps include producing the contained vent gas, Second CO 2 The steps include providing the contained vent gas to a methane-producing microbial fermentation vessel and providing organic acid to a CO fermentation vessel, Second CO 2 The process involves fermenting the contained vent gas with methane-producing archaea in a methane-producing microbial fermentation vessel to produce methane. A method that includes this.

55. Methanogenic archaea include Metanobacterium alkaliphyllum, Metanobacterium bryantyi, Metanobacterium congolense, Metanobacterium deflubyi, Metanobacterium espanolae, Metanobacterium formicicum, Metanobacterium ibanobii, Metanobacterium parstre, Metanobacterium thermagregans, Metanobacterium euriginosum, Metanobrevibacter acididurans, and Metanobrevibacter - Arboriphyllicus, Metanobrevibacter gotscharkii, Metanobrevibacter oreyae, Metanobrevibacter luminantium, Metanobrevibacter smisi, Metanobrevibacter oozei, Metanobrevibacter wolinii, Metanothermobacter marbulgensis, Metanothermobacter thermoautotrophum, Metanothermobacter thermoflexus, Metanothermobacter thermophilus, Metanothermobacter wolfei Metanotermus sochiabilis, Metanocorpus bavaricum, Metanocorpus palham, Metanocreus thikoensis, Metanocreus submarinus, Metanogenium frigidum, Metanogenium liminatans, Metanogenium marinum, Metanosarcinia acetylborans, Metanosarcinia barkerii, Metanosarcinia mazei, Metanosarcinia thermophylla, Metanomicrobium mobile, Metanocardococcus janaskii, Metanoco The method according to claim 54, selected from the group consisting of Cass aeolicus, Metanococcus maripaldis, Metanococcus van neerii, Metanococcus voltae, Metanothermococcus thermolotrophus, Metanopyrus candrelli, Metanothermobacter thermoautotrophycus, Metanocardococcus favens, Metanocardococcus indicus, Metanocardococcus infernus, and Metanocardococcus bulcanius.

56. The method according to claim 54, wherein the methanogenic archaeon is metanothermobacter thermoautotrophycus.

57. The methane-producing microbial fermentation vessel contains approximately 1 to 3 mmol of CO2. 2 / min / cell ratio CO2 2 The method according to claim 54, which provides absorption.

58. The methane-producing bacteria fermentation vessel contains approximately 3 to 12 mmol of H. 2 / min / cell-to-gram ratio H 2 The method according to claim 54, which provides absorption.

59. The method according to claim 54, wherein the methane-producing microbial fermentation vessel has a cell retention time of about 5 to about 50 hours.

60. The method according to claim 54, wherein the methane-producing microbial fermentation vessel yields a methane production capacity of approximately 1.2 to approximately 2.5 mmol of methane / minute / cell per gram.

61. The methane-producing bacteria fermentation vessel contains more than 65% CO2. 2 The method according to claim 54, which provides a conversion rate.

62. The methane-producing bacteria fermentation container is supplied with H 2 The method according to claim 54, which receives a flow.

63. Acetic acid production CO 2 Fermentation vessel, replenishment H 2 The method according to claim 54, which receives a flow.

64. CO 2 The method according to claim 54, wherein the acetic acid-producing bacteria are selected from the group consisting of Thermoanaerobacter kivui, Acetoanaerobium noterae, Acetobacterium woodii, Alkalibaculum bacchi, Acetobacterium bakii, and mixtures thereof.

65. The method according to claim 54, wherein the organic acid is one or more C1 to C10 organic acids.

66. The method according to claim 54, wherein the organic acid is acetic acid.

67. Acetic acid production CO 2 CO2 in a fermentation vessel 2 The method according to claim 54, wherein the conversion rate is controlled to 55% to 75%.

68. CO-converting acetic acid-producing bacteria include Clostridium aceticum, Clostridium acetobutylicum, Clostridium acetobutylicum P262, Clostridium autoethanogenum (DSM 19630, German DSMZ), Clostridium autoethanogenum (DSM 10061, German DSMZ), Clostridium autoethanogenum (DSM 23693, German DSMZ), Clostridium autoethanogenum (DSM 24138, German DSMZ), Clostridium carboxydivorans, Clostridium coscatii (ATCC PTA-10522), Clostridium drachei, Clostridium longudarii PETC (ATCC 49587), Clostridium longudarii ERI2 (ATCC 55380), Clostridium longudarii C-01 (ATCC The method according to claim 54, selected from the group consisting of Clostridium 55988), Clostridium lyngdaryi O-52 (ATCC 55889), Clostridium magnum, Clostridium pastelianum (DSM 525, German DSMZ), Clostridium ragsdalei P11 (ATCC BAA-622), Clostridium scatogenes, Clostridium thermoreticum, Clostridium urtense, Clostridium sticklandii, and mixtures thereof.

69. The method according to claim 54, wherein the alcohol is ethanol.

70. The method according to claim 54, wherein the CO fermentation vessel yields a CO conversion rate of 80% or more.

71. The method according to claim 54, wherein the CO fermentation vessel yields a specific alcohol production capacity of 1 gram or more of alcohol per day per cell, with a yield of 10 grams of alcohol per day per cell.

72. A fermentation liquid broth containing CO-converting acetic acid-producing bacteria is obtained from a CO-fermentation vessel, and acetic acid-producing CO is produced. 2 CO2 from the fermentation vessel 2 The steps include obtaining a fermented liquid broth containing acetic acid-producing bacteria, and obtaining a fermented liquid broth containing methane-producing archaea from a methane-producing bacteria fermentation vessel, Fermented liquid broth containing CO-converting acetic acid-producing bacteria, CO 2 The process involves separating a fermented liquid broth containing acetic acid-producing bacteria and a fermented liquid broth containing methane-producing archaea into a cell-free permeate and a cell-containing suspension. The steps include: rupturing the cell membrane of a cell-containing suspension to generate a homogenate; The steps include: using a fractionation apparatus to fractionate the homogenate into a protein-containing supernatant and a protein-containing cell debris portion; Steps to obtain protein-containing nutritional supplements and The method according to claim 54, further comprising:

73. The method according to claim 72, wherein the cell-containing suspension has a dry cell mass concentration of about 20 g / liter to about 200 g / liter.

74. The method according to claim 72, wherein the step of rupturing the cell membrane of a cell-containing suspension is performed using one or more rupture devices selected from the group consisting of microfluidic devices, sonic processing devices, ultrasonic devices, mechanical destruction devices, French presses, freezers, heaters, heat exchangers, distillation columns, sterilization devices, UV sterilization devices, gamma ray sterilization devices, reactors, homogenizers, and combinations thereof.

75. The method according to claim 72, wherein, before the step of rupturing the cell membranes of the cell-containing suspension, the pH of the cell-containing suspension is adjusted to about 6 to about 12.

76. The method according to claim 72, wherein the homogenate is a hydrolyzed lysate formed by contacting a cell-containing suspension with a hydrolase enzyme.

77. The method according to claim 76, wherein a cell-containing suspension and a hydrolase enzyme are incubated at a temperature of about 50 to about 70°C for about 3 to about 72 hours to form a hydrolyzed lysate.

78. The method according to claim 76, wherein the hydrolase enzyme is selected from the group consisting of subyrase, alcalase, serine protease, serine endopeptidase, and mixtures thereof.

79. The method according to claim 72, wherein the homogenate is fractionated into a protein-containing supernatant and a protein-containing cell debris portion using centrifugation, ultrafiltration, and / or a combination thereof.

80. The method according to claim 72, wherein the protein-containing supernatant has a nucleic acid content of less than about 5%.

81. The method according to claim 72, wherein the protein-containing supernatant is dehydrated to obtain a soluble protein-containing nutritional supplement containing about 60 to about 99% dry mass of protein.

82. CO and CO 2 A method of converting, CO and H 2 A gaseous substrate containing CO is fermented in a CO fermentation vessel with CO-converting acetic acid-producing bacteria to produce alcohol and the first CO 2 The steps include producing the contained vent gas, At least a portion of the first CO 2 The contained vent gas is converted into acetic acid CO from the CO fermentation vessel. 2 Steps for providing to the fermentation vessel, At least a portion of the first CO 2 The contained vent gas is converted into acetic acid CO2. 2 CO2 in the fermentation vessel 2 The steps include: fermenting with acetic acid-producing bacteria to produce organic acids, The steps include providing organic acids to a CO fermentation vessel, At least the first CO 2 The steps include providing another portion of the contained vent gas from the CO fermentation vessel to the methane-producing bacteria fermentation vessel, At least the first CO 2 The other part of the contained vent gas is fermented by methane-producing archaea in a methane-producing fermentation vessel to produce methane. A method that includes this.

83. Acetic acid production CO 2 The fermentation vessel is the second CO 2 Further production of contained vent gas, second CO 2 The method according to claim 82, wherein the contained vent gas is supplied to a methane-producing microbial fermentation vessel.

84. The method according to claim 82, wherein the methane-producing microbial fermentation vessel yields a methane production capacity of approximately 1.2 to approximately 2.5 mmol of methane / minute / cell per gram.

85. The methane-producing bacteria fermentation vessel contains more than 65% CO2. 2 The method according to claim 82, which provides a conversion rate.

86. Acetic acid production CO 2 CO2 in a fermentation vessel 2 The method according to claim 82, wherein the conversion rate is controlled to 55% to 75%.

87. The method according to claim 82, wherein the CO fermentation vessel yields a CO conversion rate of 80% or more.

88. The method according to claim 82, wherein the CO fermentation vessel yields a specific alcohol production capacity of 1 gram or more of alcohol per day per cell, with a yield of 10 grams of alcohol per day per cell.

89. A fermentation liquid broth containing CO-converting acetic acid-producing bacteria is obtained from a CO-fermentation vessel, and acetic acid-producing CO is produced. 2 CO2 from the fermentation vessel 2 The steps include obtaining a fermented liquid broth containing acetic acid-producing bacteria, and obtaining a fermented liquid broth containing methane-producing archaea from a methane-producing bacteria fermentation vessel, Fermented liquid broth containing CO-converting acetic acid-producing bacteria, CO 2 The process involves separating a fermented liquid broth containing acetic acid-producing bacteria and a fermented liquid broth containing methane-producing archaea into a cell-free permeate and a cell-containing suspension. The steps include: rupturing the cell membrane of a cell-containing suspension to generate a homogenate; The steps include: using a fractionation apparatus to fractionate the homogenate into a protein-containing supernatant and a protein-containing cell debris portion; Steps to obtain protein-containing nutritional supplements and The method according to claim 82, further comprising:

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