Process for bioconversion of carbon monoxide and carbon dioxide

Abstract

A method for bioconverting carbon monoxide and carbon dioxide is provided. More specifically, the method comprises fermenting a carbon monoxide and carbon dioxide containing substrate with acetogenic bacteria. The method achieves high levels of carbon monoxide and carbon dioxide conversion and hydrogen utilization.

Description

[Technical field]

[0001] This application claims the benefit of U.S. Provisional Patent Application Nos. 62 / 716,083, filed August 8, 2018, 62 / 716,071, filed August 8, 2018, 62 / 716,053, filed August 8, 2018, 62 / 741,871, filed October 5, 2018, and 62 / 741,797, filed October 5, 2018, all of which are incorporated by reference in their entireties. A method for bioconversion of carbon monoxide and carbon dioxide is provided. More specifically, the method includes feeding a gas stream containing carbon monoxide and carbon dioxide to acetogenic bacteria. The method achieves high levels of carbon monoxide and carbon dioxide conversion, and utilization of hydrogen. [Background technology]

[0002] The production of carbon dioxide and carbon monoxide occurs from natural processes and industrial processes, including the combustion of fossil fuels such as coal, oil and natural gas. Atmospheric concentrations of carbon dioxide and carbon monoxide continue to increase, due in part to industrial processes. This increase in the concentrations of carbon dioxide and carbon monoxide may contribute to atmospheric changes that result in climate change and global warming. Carbon dioxide is difficult to utilize in biological processes due to its high oxidation state. In addition to carbon dioxide and carbon monoxide, many industrial processes also result in the production of hydrogen. Hydrogen has a high level of reduction potential. However, hydrogen is difficult to store and utilize due to its highly flammable nature. In view of the large amounts of carbon dioxide and carbon monoxide being produced, there is a need for a bacterial fermentation system that is capable of utilizing both carbon monoxide and carbon dioxide. Summary of the Invention

[0003] The method includes the steps of: supplying a gaseous substrate Gx to a bioreactor Bx, the gaseous substrate Gx including CO2 and containing about 5 to about 90 mol% CO2. An acetogenic bacterium Mx is supplied to the bioreactor Bx. The acetogenic bacterium Mx includes an active sodium-transporting ATPase during fermentation in the bioreactor Bx. The method includes the steps of supplying sodium ions to the bioreactor Bx by one or more sodium ion sources, and fermenting the gaseous substrate Gx by the acetogenic bacterium Mx in a fermentation broth including the acetogenic bacterium Mx and one or more sodium ion sources to produce one or more organic acids. The fermentation broth includes less than about 0.01 grams of yeast extract per liter, less than about 0.01 grams of carbohydrate per liter, and the sodium ions are supplied at a sodium feed rate of about 290 to about 8750 μg / gram of cells / minute. The method includes the step of maintaining the fermentation broth at a pH in the range of about 4 to about 6.9. At least a portion of the one or more organic acids is fed to the bioreactor Bi. The method further comprises feeding a gaseous substrate Gi to the bioreactor Bi, the gaseous substrate Gi comprising CO and containing about 5 to about 90 mol % CO. The acetogenic bacteria Mi is fed to the bioreactor Bi. The acetogenic bacteria Mi comprises an active proton-translocating ATPase during fermentation in the bioreactor Bi. The method further comprises fermenting the gaseous substrate Gi with the acetogenic bacteria Mi in the bioreactor Bi in a fermentation broth comprising the acetogenic bacteria Mi to produce a liquid stream comprising one or more alcohols and a gas stream Gp comprising CO2. The method includes the steps of: supplying a gaseous substrate Gx to a bioreactor Bx, the gaseous substrate Gx comprising CO2 and H2, and containing about 5 to about 90 mol% CO2. The acetogenic bacteria Mx is supplied to the bioreactor Bx. The acetogenic bacteria Mx comprises an active sodium-transporting ATPase during fermentation in the bioreactor Bx. The method includes the steps of supplying sodium ions to the bioreactor Bx by one or more sodium ion sources, and fermenting the gaseous substrate Gx by the acetogenic bacteria Mx in a fermentation broth comprising the acetogenic bacteria Mx and one or more sodium ion sources to produce one or more organic acids. The fermentation broth comprises less than about 0.01 grams of yeast extract per liter, less than about 0.01 grams of carbohydrate per liter, and the sodium ions are supplied at a sodium feed rate of about 290 to about 8750 μg / gram of cells / minute. The method includes the step of maintaining the fermentation broth at a pH in the range of about 4 to about 6.9. At least a portion of the one or more organic acids is fed to the bioreactor Bi. The method further comprises feeding a gaseous substrate Gi to the bioreactor Bi, the gaseous substrate Gi comprising CO and containing about 5 to about 90 mol % CO. The acetogenic bacteria Mi is fed to the bioreactor Bi. The acetogenic bacteria Mi comprises an active proton-translocating ATPase during fermentation in the bioreactor Bi. The method further comprises fermenting the gaseous substrate Gi with the acetogenic bacteria Mi in a fermentation broth comprising the acetogenic bacteria Mi in a bioreactor system Bi-s to produce a liquid stream comprising one or more alcohols and a gas stream Gp comprising CO2. The method includes the steps of: supplying a gaseous substrate Gx to a bioreactor Bx, the gaseous substrate Gx including CO2 and containing about 5 to about 90 mol% CO2. The acetogenic bacteria Mx is supplied to the bioreactor Bx, the acetogenic bacteria Mx including an active sodium-transporting ATPase during fermentation in the bioreactor Bx. The method includes the steps of supplying sodium ions to the bioreactor Bx by one or more sodium ion sources, and fermenting the gaseous substrate Gx by the acetogenic bacteria Mx in a fermentation broth including the acetogenic bacteria Mx and one or more sodium ion sources to produce one or more organic acids. The fermentation broth includes less than about 0.01 grams of yeast extract per liter, less than about 0.01 grams of carbohydrate per liter, and the sodium ions are supplied at a sodium feed rate of about 290 to about 8750 μg / gram of cells / minute. The method includes the step of maintaining the fermentation broth at a pH in the range of about 4 to about 6.9. At least a portion of the one or more organic acids is fed to the bioreactor system Bi-s. The method further comprises feeding a gaseous substrate Gi to the bioreactor system Bi-s, the gaseous substrate Gi comprising CO and containing about 5 to about 90 mol% CO. The acetogenic bacteria Mi is fed to the bioreactor system Bi-s, the acetogenic bacteria Mi comprising an active proton-translocating ATPase during fermentation in the bioreactor system Bi-s. The method further comprises fermenting the gaseous substrate Gi by the acetogenic bacteria Mi in the bioreactor system Bi-s in a fermentation broth comprising the acetogenic bacteria Mi to produce a liquid stream comprising one or more alcohols and a gas stream Gp comprising CO2.

[0004] So that the above-recited features of the present disclosure may be understood in detail, reference may be made to the embodiments having a more particular description of the present disclosure briefly summarized above, some of which are illustrated in the accompanying drawings. It should be noted, however, that the accompanying drawings merely illustrate typical embodiments of the present disclosure and therefore should not be considered as limiting the scope of the present invention, as other equally effective embodiments may be recognized in the present disclosure. [Brief description of the drawings]

[0005] [Figure 1] FIG. 1 is a schematic diagram of a system for producing one or more oxygenated hydrocarbon compounds from a fermentation process using two bioreactors. [Diagram 2] FIG. 1 is a schematic diagram of a water recycling system. [Diagram 3] FIG. 1 is a schematic diagram of a system for producing one or more oxygenated hydrocarbon compounds from a fermentation process using multiple bioreactors. [Figure 4] 1 is a graph of the conversion of CO2 and H2 by Acetobacterium woodii in a bioreactor. [Diagram 5] 1 is a graph illustrating acetate production by Acetobacterium woodii. [Figure 6] 1 is a graph showing specific acetic acid productivity. [Figure 7] 1 is a graph showing the utilization of CO, CO2, and H2 by Acetobacterium woodii. [Figure 8] 1 is a graph illustrating CO2 conversion, H2 conversion, and cell density of Acetobacterium woodii at pH 5.2 without a chelating agent in the growth medium. [Figure 9] FIG. 1 is a graph illustrating the growth of Acetobacterium woodii using ethylenediaminediacetic acid (EDDA) as a chelating (complexing) agent in the growth medium. [Figure 10] 1 is a graph illustrating the effect of molybdenum on acetate production by Acetobacterium woodii. [Figure 11] 1 is a graph illustrating the effect of molybdenum on required gas flow rate and cell density of Acetobacterium woodii. [Figure 12]1 is a graph illustrating the specific gas uptake by Clostridium ljungdahlii with various concentrations of acetate. [Figure 13] FIG. 1 is a graph showing CO and CO2 utilization by Clostridium ljungdahlii and Acetobacterium woodii in a two-stage bioreactor system. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0006] The following description is not to be taken in a limiting sense, but is made merely for the purpose of describing the general principles of exemplary embodiments. The scope of the present disclosure should be determined with reference to the claims.

[0007] definition For the purposes of this disclosure, unless otherwise defined, the following terms used throughout the specification are defined as follows and can include either the singular or plural forms of the definitions defined below. The term "about" modifying any quantity refers to the variation of the quantity encountered in actual conditions, for example, in a laboratory, pilot plant, or production facility. For example, the amount of a component or measurement used in a mixture or amount, when modified by "about", includes the variation and degree of care that is usually taken when measuring in experimental conditions in a manufacturing plant or laboratory. For example, the amount of a component of a product, when modified by "about", includes the variation between batches in multiple experiments in a plant or laboratory, and the variation inherent in the analytical method. The above amount, whether modified by "about" or not, includes the equivalent of such amount. Any amount specified in this specification and modified by "about" can also be used in this disclosure as the amount not modified by "about". The term "fermenter" includes fermenters / bioreactors consisting of one or more vessels and / or towers, or piping, including batch reactors, semi-batch reactors, continuous reactors, continuous stirred tank reactors (CSTRs), bubble column reactors, external recycle loop reactors, internal recycle loop reactors, immobilized cell reactors (ICRs), trickle bed reactors (TBRs), moving bed biofilm reactors (MBBRs), gas lift reactors, membrane reactors such as hollow fibre membrane bioreactors (HFMBRs), static mixers, gas lift fermenters, or other vessels or other devices suitable for gas-liquid contact. The terms "fermentation," "fermentation process," or "fermentation reaction," and the like, are intended to encompass both the growth phase of the process and the product biosynthesis phase. In one aspect, fermentation refers to the conversion of CO2 to acetic acid. In another aspect, fermentation refers to the conversion of CO2 to alcohol. The term "cell density" means the mass of microbial cells per unit volume of fermentation broth, e.g., grams / liter. The term "specific CO2 uptake" refers to the amount of CO2 in millimoles consumed by a unit mass (g) of microbial cells per unit time per minute, i.e., millimoles / gram / min. The term "specific CO2 uptake" refers to the amount of CO2 in millimoles consumed by a unit mass (g) of microbial cells per unit time per minute, i.e., millimoles / gram / min. As used herein, productivity is expressed as STY. In this embodiment, alcohol productivity may be expressed as STY (space time yield), expressed as g ethanol / (L day) or g acetic acid / (L day). As used herein, "oxygenated hydrocarbon compounds" may include carbon, hydrogen and oxygen containing compounds such as, for example, ethanol and butanol.

[0008] CO / CO2 Conversion System The present disclosure provides methods, systems and compositions for producing and obtaining alcohol products and bacterial proteins derived from microbial cell biomass following an anaerobic bacterial fermentation process. In one embodiment, systems and methods are provided that improve carbon capture efficiency, reduce carbon dioxide footprint, and increase productivity of alcohol production. 1, the system may include a bioreactor Bi 120 adapted to ferment a gaseous substrate Gi 186 with acetogenic bacteria Mi. The gaseous substrate Gi 186 may include carbon monoxide (CO) and hydrogen gas (H2). In one embodiment, fermentation of gaseous substrate Gi 186 in bioreactor Bi 120 results in gas stream Gp 184. Gas stream Gp 184 includes carbon dioxide (CO2) and may include one or more gases selected from the group consisting of carbon monoxide (CO), hydrogen gas (H2), methane (CH4), nitrogen (N2), and combinations thereof. In this embodiment, gas stream Gp from bioreactor Bi 120 may include about 0.5 mol% to about 50 mol% CO, in another embodiment, about 0.5 mol% to about 40 mol%, in another embodiment, about 0.5 mol% to about 30 mol%, in another embodiment, about 0.5 mol% to about 20 mol%, and in another embodiment, about 0.5 mol% to about 10 mol% CO. Furthermore, in another embodiment, the gas stream Gp originating from the bioreactor Bi 120 may contain about 5 mol% to 100 mol% CO2, in yet another embodiment, about 5 mol% to 90 mol% CO2, in another embodiment, 5 mol% to 80 mol% CO2, in another embodiment, 5 mol% to 70 mol% CO2, in another embodiment, 5 mol% to 60 mol% CO2, in another embodiment, 5 mol% to 50 mol% CO2, in another embodiment, 5 mol% to 40 mol% CO2, in another embodiment, 5 mol% to 30 mol% CO2, in another embodiment, 5 mol% to 20 mol% CO2, and in another embodiment, 5 mol% to 10 mol% CO2. As shown in FIG. 1, the system may include a bioreactor Bx 110 adapted to ferment a gaseous substrate by acetogenic bacteria Mx. A gaseous substrate Gx is supplied to the bioreactor Bx 110 in a gas line 187. In this embodiment, a gas stream Gp 184 from the bioreactor Bi 120 may be directly supplied to the bioreactor Bx 110. In another embodiment, the system may include a gas make-up line 112 for supplying additional gaseous substrate, which is blended with the gas stream Gp 184 to provide a gas stream Gx 187 that is delivered to the bioreactor Bx 110. Waste gas from the bioreactor Bx 110 may be discharged through a vent line 207. Both bioreactors are supplied with nutrients from a nutrient supply tank 196. The nutrient supply tank 196 may include multiple subunits for supplying the same or different nutrients to each bioreactor. Additionally, the system may also include a fluid line 182 connecting the bioreactor Bx 110 to the bioreactor Bi 120 to deliver one or more acid compounds from the bioreactor 110 Bx to the bioreactor Bi 120. The one or more acid compounds produced from the bioreactor Bx 110 include C1-C10 organic acids. Examples of C1-C10 organic acids include acetic acid, formic acid, propionic acid, butyric acid, pentanoic acid (valeric acid), hexanoic acid, heptanoic acid, decanoic acid, and combinations thereof. In one embodiment, the acid compounds from the bioreactor Bx 110 delivered to the bioreactor Bi 120 are effective to increase alcohol production in the bioreactor Bi 120. In an embodiment where the organic acid is acetic acid, a gaseous substrate Gi 186 is delivered to the bioreactor Bi 120 to maintain an acetic acid concentration of about 5 g / L or less in the bioreactor Bi 120. When the gaseous substrate Gi is fed to reduce the concentration of acetate, the concentration of butyric acid also decreases. Therefore, acetate is used as an indicator to maintain an appropriate amount of organic acid in the bioreactor Bi. In another embodiment, the feed rate of organic acid is used to maintain an acetate concentration of about 5 g / L or less in the bioreactor Bi 120. 1, the cell permeate line 192 is configured to feed the permeate to a distillation column 188 for separating products 190 from the permeate. The products may include alcohol-containing products, including ethanol, butanol, and combinations thereof. Water (distillation bottoms) may be returned to bioreactor BX 110 via water return line 202 and / or to bioreactor Bi 120 via water return line 203.

[0009] FIG. 2 illustrates a more detailed embodiment of the water recycle system. In this embodiment, the organic acids produced in the bioreactor Bx 110 are conveyed through the microfiltration 314 and further fed to the bioreactor Bi 120 through the fluid line 182. At least a portion of the cells may be returned from the microfiltration 314 to the bioreactor Bx 110 by the cell recycle line 318. The broth from the bioreactor Bi 120 is conveyed through the line 194 to the microfiltration 301. At least a portion of the cells may be returned from the microfiltration 301 to the bioreactor Bi 120 by the cell recycle line 317. In some optional embodiments, the liquid from the line 194 can be directed to the ultrafiltration 307, and at least a portion of the cells may be returned from the ultrafiltration 307 to the bioreactor Bi 120. The liquid from the ultrafiltration 307 is sent to the distillation feed tank 311 and then to the distillation column 188. Water (distillation bottoms) may be returned to bioreactor BX 110 via water return line 202 and / or to bioreactor Bi 120 via water return line 203.

[0010] In another embodiment, as shown in Figure 3, the method may include a bioreactor Bx 110 adapted to ferment a gaseous substrate Gx 187 with acetogenic bacteria Mx. The gaseous substrate Gx 187 may include carbon monoxide (CO) and hydrogen gas (H2) in addition to CO2. As shown in Figure 3, the gaseous substrate Gx may be supplied to the bioreactor Bx from a bioreactor system Bi-s, which includes two or more bioreactors Bi-n. As shown in Figure 3, Bi-n includes two bioreactors Bi 120.

[0011] The gaseous substrate Gx 187 is supplied to the bioreactor Bx 110 in one or more gas lines. In this embodiment, the gas stream Gp 184 from two or more bioreactors Bi 120 of the bioreactor system Bi-s can be directly supplied to the bioreactor Bx 110. In another embodiment, the system can include a gas make-up line 112 for supplying additional gaseous substrate, which is blended with the gas stream Gp 184 to provide the gas stream Gx 187 that is delivered to the bioreactor Bx 110. The waste gas from the bioreactor Bx 110 can be discharged from a vent line 207. Each bioreactor can be supplied with nutrients from one or more nutrients supply tanks 196. The nutrient supply tank 196 can include multiple subunits for supplying the same or different nutrients to each bioreactor.

[0012] The bioreactor system Bi-s may include two or more bioreactors Bi 120 adapted to ferment a gaseous substrate Gi 186 by the acetogenic bacteria Mi. The gaseous substrate Gi 186 may include carbon monoxide (CO) and hydrogen gas (H2). Fermentation of the gaseous substrate Gi 186 in the bioreactor Bi 120 of the bioreactor system Bi-s results in one or more gas streams Gp 184. The gas streams Gp 184 include carbon dioxide (CO2) and may include one or more gases selected from the group consisting of carbon monoxide (CO), hydrogen gas (H2), methane (CH4), nitrogen (N2) and combinations thereof. In this embodiment, the gas stream Gp from the bioreactor Bi 120 may contain about 0.5 mol % to about 50 mol % CO, in another embodiment, about 0.5 mol % to about 40 mol %, in another embodiment, about 0.5 mol % to about 30 mol %, in another embodiment, about 0.5 mol % to about 20 mol %, and in another embodiment, about 0.5 mol % to about 10 mol % CO. Furthermore, in another embodiment, the gas stream Gp from the bioreactor Bi 120 may contain about 5 mol% to 100 mol% CO2, in yet another embodiment, about 5 mol% to 90 mol% CO2, in another embodiment, 5 mol% to 80 mol% CO2, in another embodiment, 5 mol% to 70 mol% CO2, in another embodiment, 5 mol% to 60 mol% CO2, in another embodiment, 5 mol% to 50 mol% CO2, in another embodiment, 5 mol% to 40 mol% CO2, in another embodiment, 5 mol% to 30 mol% CO2, in another embodiment, 5 mol% to 20 mol% CO2, and in another embodiment, 5 mol% to 10 mol% CO2. In one embodiment, gaseous substrate Gi 186 is fed to two or more bioreactors Bi 120 (collectively referred to as Bi-n) to achieve a gas superficial velocity effective to produce about 10% or less of the culture volume per hour of foam.

[0013] Additionally, the system may also include two or more fluid lines 182 connecting the bioreactor Bx 110 to the bioreactor system Bi-s to feed one or more acid compounds from the bioreactor Bx 110 to the bioreactor system Bi-s. In this embodiment, the bioreactor system Bi-s may include two or more bioreactors Bi 120 (two bioreactors Bi 120 are shown). The one or more acid compounds produced from the bioreactor Bx 110 include C1-C10 organic acids. Examples of C1-C10 organic acids include acetic acid, formic acid, propionic acid, butyric acid, pentanoic acid (valeric acid), hexanoic acid, heptanoic acid, decanoic acid, and combinations thereof. In one embodiment, the acid compounds from the bioreactor Bx 110 fed to the bioreactor system Bi-s are effective to increase alcohol production in the bioreactor system Bi-s.

[0014] 3, the cell permeate line 192 is configured to feed the permeate to a distillation column 188 for separating products 190. The products may include alcohol-containing products, including ethanol, butanol, and combinations thereof. Water (distillation bottoms) may be returned to bioreactor Bx 110 via water return line 202 and / or to bioreactor Bi 120 via water return line 203.

[0015] Bioreactor for CO conversion CO-containing substrate: The CO-containing substrate (referred to as gaseous substrate Gi 186) may include any gas that contains CO. In this embodiment, the CO-containing gas may include synthesis gas, industrial gas, and mixtures thereof. In a related embodiment, the gaseous substrate supplied to the bioreactor Bi 120 may include, in addition to CO, nitrogen gas (N2), carbon dioxide (CO2), methane gas (CH4), synthesis gas, and combinations thereof. The synthesis gas may be provided from any known source. In one embodiment, the synthesis gas may be sourced from gasification of carbonaceous materials. Gasification involves partial combustion of biomass with limited oxygen supply. The resulting gas may include CO and H2. In this embodiment, the synthesis gas will contain at least about 10 mol% CO, in one embodiment at least about 20 mol%, in one embodiment about 10 to about 100 mol%, 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. Some examples of suitable gasification methods and apparatus are provided in U.S. Serial Nos. 61 / 516,667, 61 / 516,704, and 61 / 516,646, all of which were filed on April 6, 2011, and U.S. Serial Nos. 13 / 427,144, 13 / 427,193, and 13 / 427,247, all of which were filed on March 22, 2012, all of which are incorporated herein by reference. In another embodiment, the method has applicability in aiding the production of alcohol from gaseous substrates, such as industrial gases that contain a high volume of CO. In some embodiments, the gas containing CO comes from carbon-containing waste, such as industrial waste gases, or from the gasification of other wastes. Thus, the method is an effective way to capture carbon that would otherwise be emitted into the environment. Examples of industrial gases include gases produced during steel manufacturing, non-ferrous manufacturing, oil refining processes, coal gasification, biomass gasification, power generation, carbon black production, ammonia production, methanol production, coke manufacturing, and gas reforming. In another embodiment, H2 may be supplied from industrial waste gases or from gasification of other waste materials. Thus, the method is an effective way to capture hydrogen that would otherwise be discharged to the environment. Examples of industrial gases include gases produced during steel manufacturing, non-ferrous manufacturing, petroleum refining processes, coal gasification, biomass gasification, power generation, carbon black production, ammonia production, methanol production, and coke production. Other sources of hydrogen may include, for example, electrolysis of H2O and biogenic H2. The CO-containing substrate may be fed directly to the fermentation process or may be further modified to contain an appropriate H2 to CO molar ratio, depending on the composition of the CO-containing substrate. In one embodiment, the CO-containing substrate fed to the fermentor has a molar ratio of H2 to CO of about 0.2 or greater, in another embodiment about 0.25 or greater, and in another embodiment about 0.5 or greater. In another embodiment, the CO-containing substrate fed to the fermentor may contain about 40 molar percent CO and H2 or greater, and about 30 molar percent CO or less, in another embodiment about 50 molar percent CO and H2 or greater, and about 35 molar percent CO or less, and in another embodiment about 80 molar percent CO and H2 or greater, and about 20 molar percent CO or less. In one embodiment, the CO-containing substrate comprises CO and H2. In this embodiment, the CO-containing substrate will contain at least about 10 mol% CO, in one embodiment at least about 20 mol%, in one embodiment about 10 to about 100 mol%, 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 one embodiment, the gas separator is configured to substantially separate at least a portion of the gas stream, the portion including one or more components. For example, the gas separator can separate CO2 from a gas stream including the following components: CO, CO2, H2, and the CO2 may be passed to a CO2 storage unit and the remainder of the gas stream (including CO and H2) may be passed to a bioreactor. Any gas separator known in the art may be utilized. In this embodiment, the syngas fed to the fermenter will have about 10 mol% or less CO2, in another embodiment about 1 mol% or less CO2, and in another embodiment about 0.1 mol% or less CO2. A particular gas stream may contain a high concentration of CO and a low concentration of H. In one embodiment, it may be desirable to optimize the composition of the substrate stream to achieve more efficient alcohol production and / or total carbon capture. In another embodiment, the substrate stream is passed through a bioreactor after the substrate stream has been enriched in H. According to certain embodiments of the present disclosure, streams from two or more sources can be combined and / or blended to produce a desired and / or optimized substrate stream. For example, a stream containing high concentrations of CO, such as exhaust gas from a steel mill converter, can be combined with a stream containing high concentrations of H, such as exhaust gas from a steel mill coke oven. Depending on its composition, it may also be desirable to treat the gaseous CO-containing substrate to remove any undesirable impurities, such as dust particles, prior to introduction into the fermentation For example, the gaseous substrate may be filtered or scrubbed using known methods.

[0016] Bioreactor Design and Operation: A description of fermentor design is provided in U.S. Serial Nos. 13 / 471,827 and 13 / 471,858, both filed May 15, 2012, and U.S. Serial No. 13 / 473,167, filed May 16, 2012, all of which are incorporated herein by reference. In one embodiment, the fermentation process is initiated by the addition of a medium to the reactor. Some examples of medium compositions are described in U.S. Serial Nos. 61 / 650,098 and 61 / 650,093, filed May 22, 2012, and U.S. Patent No. 7,285,402, filed July 23, 2001, all of which are incorporated herein by reference. The medium may be sterilized to remove undesirable microorganisms, and the reactor is inoculated with the desired microorganisms. Sterilization may not be necessary.

[0017] The concentrations of the various medium components for use in bioreactor Bi 120 with acetogenic bacteria Mi are as follows: [Table 1]

[0018] The ability of certain acetogens to utilize CO is due in part to the presence of a proton or hydrogen pump, also referred to as a proton-transporting ATPase. Both proton-transporting ATPases and 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. The term proton-transporting ATPase may be used interchangeably with the term proton-dependent ATPase, and the term sodium-transporting ATPase may be used interchangeably with the term sodium-dependent ATPase. Thus, in one aspect, the method includes carrying out fermentation in a fermentation bioreactor including the acetogenic bacterium Mi, which includes a proton-transporting ATPase.Examples of useful acetogenic bacteria Mi include strains of Clostridium ljungdahlii, including those described in WO2000 / 68407, EP117309, U.S. Pat. Nos. 5,173,429, 5,593,886 and 6,368,819, WO1998 / 00558 and WO2002 / 08438, Clostridium autoethanogenum, including those described in WO2007 / 117157 and WO2009 / 151342 (DSM10061 and DSM19630 of DSMZ, Germany), and the strains of acetogenic bacteria Mi, respectively, described in U.S. Pat. No. 7,704,723 and in Hasan Atiyeh, "Biofuels and bioproducts from Suitable microorganisms include those of the genus Clostridium, such as strains of Clostridium ragsdalei (P11, ATCC BAA-622) and Alkalibaculum bacchi (CP11, ATCC BAA-1772), including those described in "Biomass-Generated Synthesis Gas," and Clostridium carboxidivorans (ATCC PTA-7827), as described in U.S. Patent Application Publication No. 2007 / 0276447. Other suitable microorganisms include those of the genera Moorella, including Moorella sp. HUC22-1, and Carboxydothermus. All of these references are incorporated herein by reference. Mixed cultures of two or more microorganisms may be used.

[0019] Additional examples of useful acetogenic bacteria Mi include Acetogenium kivui, Acetoanaerobium noterae, Acetobacterium woodii, Alkalibacrum bacchii CP11 (ATCC BAA-1772), Blautia producta, Butyribacterium methylotrophicum, Caldanaerobacter subterraneous, Caldanaerobacter subterraneous pacificus, Carboxydothermus hydrogenoformans, Clostridium aceticum, and others. aceticum), Clostridium acetobutylicum, Clostridium acetobutylicum P262 (DSM19630, DSMZ, Germany), Clostridium autoethanogenum (DSM19630, DSMZ, Germany), Clostridium autoethanogenum (DSM10061, DSMZ, Germany), Clostridium autoethanogenum (DSM23693, DSMZ, Germany), Clostridium autoethanogenum (DSM24138, DSMZ, Germany), Clostridium carboxydivorans P7 (ATCC PTA-7827), 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 pasteurianumpasteurianum (DSM525 from DSMZ, Germany), Clostridium ragsdalei P11 (ATCC BAA-622), Clostridium scatologenes, Clostridium thermoaceticum, Clostridium ultunense, Desulfotomaculum kuznetsovii, Eubacterium limosum, Geobacter sulfurreducens, Methanosarcina acetivorans, Methanosarcina barkeri, Moorella thermoacetica, thermoacetica, Moorella thermoautotrophica, Oxobacter pfennigii, Peptostreptococcus productus, Ruminococcus productus, Thermoanaerobacter kivui, Clostridium stick-landii and mixtures thereof. Fermentation should desirably be carried out under appropriate conditions (e.g., CO to ethanol) for the desired fermentation to occur. Reaction conditions to be considered include pressure, temperature, gas flow rate, liquid flow rate, medium pH, medium redox potential, agitation rate (if a stirred tank reactor is used), inoculum level, maximum gas substrate concentration to ensure that CO in the liquid phase is not rate limiting, and maximum product concentration to avoid product inhibition.

[0020] Start-up: Upon inoculation, an initial feed gas rate effective to feed an initial population of microorganisms is established. The effluent gas is analyzed to determine the content of the effluent gas. The results of the gas analysis are used to control the rate of the feed gas. In this embodiment, the method results in a ratio of calculated amount of CO (mmoles) to initial cell density (grams / liter) that is about 0.2 to about 0.9, in another embodiment, about 0.5 to about 0.9, in another embodiment, about 0.6 to about 0.8, in another embodiment, about 0.5 to about 0.7, in another embodiment, about 0.2 to about 0.5, and in another embodiment, about 0.5 to about 0.6. In another embodiment, the fermentation process includes providing syngas to the fermentation medium in an amount effective to achieve a calculated initial CO concentration in the fermentation broth of about 0.05 mM to about 0.10 mM, in another embodiment about 0.15 mM to about 0.50 mM, in another embodiment about 0.15 mM to about 0.35 mM, in another embodiment about 0.20 mM to about 0.30 mM, and in another embodiment about 0.23 mM to about 0.27 mM. Dissolved CO was calculated using Henry's law and the kLa of the reactor. The method is effective in increasing cell density as compared to the starting cell density. As used herein, the target cell density refers to a cell density of about 2.0 grams / liter or more, in another embodiment, about 2 to about 30 grams / liter, in another embodiment, about 2 to about 25 grams / liter, in another embodiment, about 2 to about 20 grams / liter, in another embodiment, about 2 to about 10 grams / liter, in another embodiment, about 2 to about 8 grams / liter, in another embodiment, about 3 to about 30 grams / liter, in another embodiment, about 3 to about 6 grams / liter, and in another embodiment, about 4 to about 5 grams / liter.

[0021] After start-up: Once the desired level is reached, the liquid phase and cell material are withdrawn from the reactor and the medium is replenished. After start-up, the cell density will be maintained at a constant level.

[0022] Bioreactor Bx for converting CO2 CO2-containing substrate: In one embodiment, the method includes feeding a CO2-containing gaseous substrate (described as gaseous substrate Gx 187) to a bioreactor. The CO2-containing substrate may include any gas that contains CO2. In this embodiment, the CO2-containing gas may include industrial gases, such as fermenter gas streams, including fermenter exhaust gases, and mixtures thereof. In a related embodiment, the CO2-containing substrate may include hydrogen or may be blended with a hydrogen source to provide the desired level and ratio of H2 to CO2.

[0023] Industrial gas: In one aspect, the method includes feeding a C02-containing substrate to a bioreactor, where the C02-containing substrate is produced from an industrial gas. Some examples of industrial gases include steel mill gas, industrial gas, and incinerator exhaust gas. Examples of industrial gases include gases produced during steel manufacturing, non-ferrous manufacturing, petroleum refining processes, coal gasification, biomass gasification, power generation, carbon black production, ammonia production, methanol production, and coke production. Hydrogen sources may include fossil fuels, steam reforming, methane oxidation, coal gasification, and water electrolysis. Depending on its composition, it may also be desirable to treat the gaseous CO2-containing substrate to remove any undesirable impurities, such as dust particles, prior to introduction into the fermentation. For example, the gaseous substrate may be filtered or scrubbed using known methods. Furthermore, depending on the composition of the CO2-containing substrate, the method may include adjusting the CO2-containing substrate to increase or decrease the concentration of CO2 and / or H2 to fall within a desired range.

[0024] Fermenter Gas Stream: In one aspect, the method includes feeding a CO2-containing substrate to a bioreactor, where the CO2-containing substrate becomes the fermenter gas stream. Some examples of fermenter gas streams include fermenter off-gas produced during syngas fermentation. Some examples of syngas fermentation are described in U.S. Patent No. 7,285,402, filed July 23, 2001, which is incorporated herein by reference. In one embodiment, the method has applicability in aiding the production of alcohol from gaseous substrates, such as industrial gases that contain high volumes of CO. In some embodiments, the gas containing CO originates from carbon-containing waste, such as industrial waste gases, or from gasification of other waste materials. Fermentation of the CO-containing gas can result in CO2 in the exhaust gas of the fermenter. Thus, the method is an effective way to capture carbon that would otherwise be emitted to the environment. In this embodiment, the exhaust gas from the fermentation of the CO-containing gas can contain about 0.5 mol% to about 50 mol% CO.

[0025] Blending of Gas Streams: According to certain embodiments, streams from two or more sources can be combined and / or blended to produce a desired and / or optimized substrate stream. For example, a stream containing high concentrations of CO2, such as exhaust gas from a steel mill, can be combined with a stream containing high concentrations of H2, such as exhaust gas from a steel mill coke oven. The CO2-containing raw substrate may be fed directly to the fermentation process or may be further modified to contain an appropriate H2 to CO2 molar ratio, depending on the composition of the CO2-containing substrate. The CO2-containing substrate may contain about 5 to about 90 mol% CO2 and about 5 to about 90 mol% H2. In one embodiment, the CO2-containing gas stream contains about 5 to about 66.6% CO2. In another embodiment, the CO2-containing substrate supplied to bioreactor Bx 110 may contain about 0 mol % to about 50 mol % CO, in another embodiment, about 0.5 mol % CO to about 50 mol % CO, in another embodiment, about 0.5 mol % CO to about 5 mol % CO, and in another embodiment, about 2 mol % CO to about 5 mol % CO. In one embodiment, the acetogenic bacteria will have a molar consumption ratio of H2 to CO2 at a ratio of about 4:1 to about 1:2. Thus, any substrate gas, including H2 and CO2, fed to the bioreactor may be utilized. However, the optimal level of substrate gas fed to the bioreactor will have a ratio of H2 to CO2 of about 4:1 to about 1:1, in another embodiment about 2:1, and in another embodiment about 3.5:1 to about 1.5:1.

[0026] Bioreactor Design and Operation: A description of fermentor design is provided in U.S. Serial Nos. 13 / 471,827 and 13 / 471,858, both filed May 15, 2012, and U.S. Serial No. 13 / 473,167, filed May 16, 2012, all of which are incorporated herein by reference. Fermentation should desirably be carried out under appropriate conditions (e.g., CO2 vs. acetic acid) for the desired fermentation to occur. Reaction conditions to be considered include pressure, temperature, gas flow rate, liquid flow rate, medium pH, agitation rate (if a stirred tank reactor is used), inoculum level, and maximum acetic acid concentration to avoid product inhibition. In this embodiment, the method includes the following ranges of reaction conditions: Pressure: from about 0 to about 500 psi, in another embodiment, from about 0 to about 200 psi; Temperature: about 30℃~about 42℃; pH of the medium: about 4 to about 6.9; Stirring speed: about 100 to about 2000 rpm; Provision of nutrients as described herein.

[0027] Acetogenic bacteria: In one embodiment, the microorganisms utilized include the anaerobic acetogenic bacteria Mx, which contain a sodium pump, which may also be described (in the case of membrane bioenergetics) as a sodium-transporting ATPase. The sodium-transporting ATPase is 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. Acetogens that contain a sodium-transporting ATPase require about 500 ppm NaCl in their growth medium for growth. To determine whether an acetogen contains a sodium-transporting ATPase, the acetogen is inoculated into a serum bottle containing about 30 to about 50 ml of growth medium containing about 0 to about 2000 ppm NaCl. Normal growth at a NaCl concentration of about 500 ppm or more means that the acetogen contains a sodium-transporting ATPase. In this embodiment, suitable acetogenic bacteria Mx include Acetobacterium genus bacteria, Acetogenium kibui, Acetoanaerobium notelae, Acetobacterium woodii, Alkalibacrum bacchii CP11 (ATCC BAA-1772), Moorella thermoacetica, Moorella thermoautotrophica, Ruminococcus productus, Acetogenium kibui, and combinations thereof. In another embodiment, the microorganism is Acetobacterium woodii.

[0028] Control of medium composition and medium feed rate: According to one embodiment, the fermentation process is initiated by the addition of a suitable medium to the reaction vessel. The liquid contained in the reaction vessel may comprise any type of suitable nutrient or fermentation medium. The nutrient medium will contain vitamins and minerals effective to allow growth of the microorganism used. Sterilization may not always be necessary.

[0029] The concentrations of the various media components for use in bioreactor Bx 110 with acetogenic bacteria Mx are as follows: [Table 2]

[0030] The process operation maintains a pH in the range of about 4 to about 6.9, in another embodiment about 5 to about 6.5, in another embodiment about 5.1 to about 6, and in another embodiment about 5.2 to about 6. The medium comprises less than about 0.01 g / L yeast extract and less than about 0.01 g / L carbohydrate. The composition also includes a sodium ion concentration of about 40 to about 500 mmol per liter, in another embodiment, about 40 to about 250 mmol per liter, and in another embodiment, about 50 to about 200 mmol per 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 Na, and in another embodiment, about 3000 to about 4000 ppm Na. The sodium ion source is provided by a compound selected from the group consisting of sodium chloride, sodium hydroxide, sodium phosphate, sodium sulfate, sodium nitrate, sodium bicarbonate, sodium bisulfate, and mixtures thereof. The composition includes a molybdenum source. In this embodiment, the molybdenum concentration is from about 3.97 μg / L to about 396.5 μg / L, and in another embodiment, from about 7.93 μg / L to about 198.25 μg / L. The molybdenum source can include Na2MoO4, CaMoO4, FeMoO4, and mixtures thereof. The composition may also include a complexing agent. In this embodiment, when the composition has a pH of about 5.2 or more, a complexing agent may be included in the composition. The complexing agent may include ethylenediaminetetraacetic acid (EDTA), ethylenediaminediacetic acid (EDDA), ethylenediaminedisuccinic acid (EDDS) and mixtures thereof. The composition is+ , P, K, Fe, Ni, Co, Se, Zn or Mg. The source of each of these elements may be as follows: NH4 + The nitrogen may be supplied from a nitrogen source selected from the group consisting of ammonium hydroxide, ammonium chloride, ammonium phosphate, ammonium sulfate, ammonium nitrate, and mixtures thereof. P: Phosphorus may be provided from a phosphorus source selected from the group consisting of phosphoric acid, ammonium phosphate, potassium phosphate, and mixtures thereof. K: Potassium may be provided from a potassium source selected from the group consisting of potassium chloride, potassium phosphate, potassium nitrate, potassium sulfate, and mixtures thereof. Fe: Iron may be provided from an iron source selected from the group consisting of ferrous chloride, ferrous sulfate, and mixtures thereof. Ni: Nickel may be provided from a nickel source selected from the group consisting of nickel chloride, nickel sulfate, nickel nitrate, and mixtures thereof. Co: Cobalt may be provided from a cobalt source selected from the group consisting of cobalt chloride, cobalt fluoride, cobalt bromide, cobalt iodide, and mixtures thereof. Se: Selenium is Na2SeO 3、 It can be provided by C3H6NO2Se and mixtures thereof. Zn: Zinc can be sourced from ZnSO4. W: Tungsten may be provided from a tungsten source selected from the group consisting of sodium tungstate, calcium tungstate, potassium tungstate, and mixtures thereof. Mg: Magnesium may be supplied from a magnesium source selected from the group consisting of magnesium chloride, magnesium sulfate, magnesium phosphate and mixtures thereof. S: The composition may also include sulfur. The sulfur may be provided from a sulfur source selected from the group consisting of cysteine, sodium sulfide, NaHS, NaH2S, and mixtures thereof.

[0031] Start-up: Upon inoculation, an initial feed gas rate is established effective to supply an initial population of microorganisms. The effluent gas is analyzed to determine the content of the effluent gas. The results of the gas analysis are used to control the rate of the feed gas. In this embodiment, the method results in a minimum cell density of about 0.1 grams per liter. In another embodiment, the method results in a ratio of calculated CO2 (mmol / min) per unit time to initial cell density (grams / liter) of about 0.05 to about 1, and in another embodiment, about 0.01 to about 4. In one aspect, nutrients can be added to the culture to increase the cell growth rate. Suitable nutrients may include the non-carbohydrate fraction of yeast extract.

[0032] After start-up: Once the desired level is reached, the liquid phase and cellular material are withdrawn from the reactor and the medium is replenished. The fermentation process is effective to increase the cell density compared to the starting cell density. In this embodiment, the method achieves an average cell density of about 2 to about 50 grams / liter, in another embodiment about 2 to about 30 grams / liter, in another embodiment about 2 to about 20 grams / liter, in another embodiment about 2 to about 10 grams / liter, in another embodiment about 5 to about 10 grams / liter, and in another embodiment about 2 to about 6 grams / liter. Production of organic acids: In another embodiment, the method results in a source of C1-C10 organic acids. In this embodiment, the method may include a step of obtaining an acid product or product from the fermentation liquid broth. In this embodiment, an organic acid specific productivity of about 0.2 to about 50 grams of organic acid / liter / day / g cells, in another embodiment, about 0.2 to about 20 grams of organic acid / liter / day / g cells, in another embodiment, about 10 to about 50 grams of organic acid / liter / day / g cells, in another embodiment, about 14 to about 30 grams of organic acid / liter / day / g cells, in another embodiment, about 2 to about 20 grams of organic acid / liter / day / g cells, and in another embodiment, about 15 to about 25 grams of organic acid / liter / day / g cells is achieved. In one embodiment, the organic acid is acetic acid, or butyric acid, or a mixture of both. CO2 and H2 Conversion: The method is effective to achieve a CO2 uptake of about 0.05 to about 1.5 mmol CO2 / min / gram dry cells, and a H2 uptake of about 0.08 to about 1.5 mmol H2 / min / gram dry cells. The method is effective to achieve a CO2 conversion rate of about 25 to about 100%, in another embodiment, a CO2 conversion rate of about 50 to about 100%, and in another embodiment, a CO2 conversion rate of about 75 to about 100%. In another embodiment, the method is effective to achieve a H2 conversion rate of about 25 to about 100%, in another embodiment, a H2 conversion rate of about 50 to about 100%, and in another embodiment, a H2 conversion rate of about 75 to about 100%.

[0033] 4 shows a graph of CO2 conversion 404 and H2 conversion 402 by Acetobacterium woodii. A graphical illustration of acetate production 504, and its running average 502, and cell density 506 over time is shown in FIG. EXAMPLES

[0034] Example 1 Preparation of Acetobacterium woodii The first freeze-dried pellets of Acetobacterium woodii were obtained from the German Culture Collection DSMZ, strain ID DSM-1030. Cultures were initially obtained from freeze-dried pellets using rich medium (fructose and yeast extract). An acclimation method was used to remove fructose from serum bottle medium, where the concentration of fructose in the growth medium was stepwise reduced to 75%, 50% and 10%. Growth rate and gas utilization were used as indicators of acclimation (approximately 5 weeks). A pre-pH acclimation work in serum bottles reduced the required pH from 7.4 to 6.0 (3 weeks). At this point the culture was amplified and inoculated into a reactor. In the reactor the culture was further acclimated and grown at a lower pH of 5.2-5.7.

[0035] Example 2 Start-up method of CSTR reactor with Acetobacterium woodii Syngas containing CO2 and H2 was continuously introduced into a stirred tank bioreactor containing Acetobacterium woodii along with a liquid medium containing vitamins, trace metals, cysteine ​​(as a sulfur source) and salts, as described herein. A New Brunswick Bioflow 310 reactor containing fermentation medium with actively growing Acetobacterium woodii was started. The agitation rate of the reactor was set at 200 rpm. This agitation rate was increased from 200 to 600 rpm throughout the experiment. The feed gas flow rate to the reactor was increased from an initial value of 49 mL / min to 137 mL / min. The temperature in the bioreactor was maintained at 33.5°C throughout the experiment. Samples of the gas feed to the bioreactor, and the exhaust gas from the bioreactor and the fermentation broth in the bioreactor were taken at regular intervals, e.g., the feed gas, exhaust gas and fermentation broth were sampled approximately every day, once every 2 hours and once every 4 hours. The above samples were analyzed for consumption or production of various gas components, acetate concentration of the broth and optical density (cell density) of the culture. The unusable volume of the reactor was maintained between 1600-1750 ml throughout the experiment. Similarly, the gas flow rate to the reactor was maintained at the required gas flow rate by using mass flow controllers. The feed syngas composition was 70% H2, 25% CO2 and 5% N2. The experiment was concluded when stable operation was reached. The experiment was started after the cell recycle system (CRS) was attached to the reactor. During the experiment, the flow rate of nutrients (growth medium) into the reactor was as shown in the table. The medium feed rate was maintained throughout the experiment. The feed rate of base (0.5 M NaOH) for pH control was 0.14-0.44 ml / min and 5.1-5.4 mL / min of permeate was withdrawn from the reactor by the CRS. H2 and CO2 in the feed gas were fixed in cell material and acetate. Removal of H2 and CO2 was calculated by comparing the inlet gas composition with the outlet gas composition. The uptake of the constituent gases is expressed as % of the gas molecules converted by the bacteria. In this experiment, the following conversion rates were achieved; H2: 40%-54%, CO2: 28%-70%. In this experiment, the rate of acetate production was 5-23 g / l / day.

[0036] The results can be summarized as follows: [Table 3]

[0037] Example 3 Fermentation of CO2, CO and H2 by Acetobacterium woodii Gas containing CO2 and H2 was continuously introduced into a stirred tank bioreactor containing Acetobacterium woodii along with a conventional liquid medium containing vitamins, trace metals and salts. Fermentation was initiated as described in Example 2 and then continued until stable operation. In this example, the feed gas contained 5 mol% CO. Figures 6 and 7 illustrate the growth of Acetobacterium woodii in the presence of 5% CO. Figure 6 illustrates cell density 602 and specific acetate productivity 604 versus time. Figure 7 illustrates H2 conversion 702, CO conversion 704, CO2 conversion 706, and cell density 708.

[0038] Example 4 Growth and maintenance of Acetobacterium woodii cultures at pH 5.2 without a chelating agent (EDTA) in the growth medium A gas stream containing CO2 and H2 was continuously introduced into a stirred tank bioreactor containing Acetobacterium woodii along with the growth medium described herein. A New Brunswick Bioflow 115 reactor containing fermentation medium with actively growing Acetobacterium woodii (AW) was started. The agitation rate of the reactor was set at 600 rpm. This agitation rate was maintained constant throughout the experiment. The feed gas flow rate to the reactor was maintained between 36.6 mL / min and 44.4 mL / min. The temperature in the bioreactor was maintained at 33°C throughout the experiment. The Na+ level was maintained between 3500 and 4000 ppm. Samples of the gas feed to the bioreactor, and the exhaust gas from the bioreactor, and the fermentation broth in the bioreactor were taken at regular intervals, e.g., the feed gas, exhaust gas, and fermentation broth were sampled approximately every day, once every 2 hours, and once every 4 hours, respectively. The above samples were analyzed for consumption or production of various gas components, acetate concentration of the broth, and optical density (cell density) of the culture. The unused volume of the reactor was maintained between 1900 and 2275 ml throughout the experiment. Similarly, the gas flow rate to the reactor was maintained at the required gas flow rate using a mass flow controller. The feed syngas composition for this experiment was 70% H2, 25% CO2 and 5% N2. The experiment was started after the reactor was fitted with a cell recycle system (CRS). During the experiment, the flow rate of nutrients (growth medium) into the reactor was maintained at 2.8 ml / min. The medium feed rate was maintained throughout the experiment. The average rate of base (NaOH) requirement to maintain the pH at 5.2 was 0.075 ml / min, and 2.9 ml / min of permeate was withdrawn from the reactor by the CRS. H2 and CO2 in the feed gas were fixed in cell material and acetate. H2 and CO2 removal was calculated by comparing the inlet gas composition with the outlet gas composition. The uptake of the constituent gases can be expressed as the % of gas molecules converted by the bacteria.

[0039] The following conversion rates were achieved: H2: 28%~54%. CO2: 40%~59%. The rate of acetic acid production was 0.7949 (g / L / day). The average cell density of the cultures was 1.9 g / L. The CO2 conversion 802, H2 conversion 804 and cell density 806 are shown in FIG.

[0040] Example 5 Use of EDDA in growth media Fermentation was initiated as described in Example 2, including the use of ethylenediaminediacetic acid (EDDA) as a chelating (complexing) agent. The solubility of some of the metals used in the AW medium decreases with increasing pH, so a chelating agent is used to keep the metals in solution. Once the pH of the reactor broth exceeds pH 5.2, a chelating agent is used to provide sufficient nutrients to the AW. Figure 9 shows a representative 96 hour experiment, illustrating that it is possible to maintain cell density 902 while producing an increased concentration of acetate 904.

[0041] Example 6 Effect of molybdenum removal and re-feeding on cell metabolism Fermentation was initiated as described in Example 2 and then continued until stable operation. Molybdenum was removed from the culture medium and then re-added to the growth medium after acetate productivity had fallen to 75% of its starting concentration. 10 illustrates acetate productivity 903 plotted against the flow rate 905 of the medium, with the red line indicating removal of molybdenum from the growth medium and re-addition of molybdenum to the growth medium. Starting at about 810 cumulative hours, a downward trend in HAc was observed, with removal of molybdenum occurring at about 795 cumulative hours. This downward trend decreased, plateaued, and then reversed to an upward trend at about 900 hours in response to re-addition of molybdenum to the medium. FIG. 11 illustrates cell density 906 and gas flow rate (GFR) 901 plotted against time, with the red line indicating removal of molybdenum from the growth medium and re-addition of molybdenum to the growth medium. Beginning at about 840 cumulative hours, the GFR required was less than before molybdenum removal, which occurred at about 795 cumulative hours. This downward trend reversed to an upward trend in response to the addition of molybdenum back to the medium at about 900 hours. The required gas flow rate was determined by the CO2 and H2 conversion rates of the culture.

[0042] Example 7 A continuous method for the conversion of CO using Clostridium ljungdahlii Syngas or waste gas containing CO and / or CO2 / H2 was continuously introduced into a stirred tank bioreactor containing a strain of Clostridium ljungdahlii along with a fermentation medium containing vitamins, trace metals and salts as described herein. A New Brunswick BioFlow310 reactor containing a suitable medium was inoculated with actively growing Clostridium ljungdahlii at 0.38 g / l. Prior to inoculation, the reactor agitation speed was set at 800 rpm, the gas flow rate to the reactor was adjusted to 20 ml / min, and a cell recycle system was attached to the reactor. At intervals of 1-4 hours, gas and liquid samples taken from the reactor were analyzed for consumption or production of various gas components, acetate concentration in the broth, ethanol concentration in the broth, and optical density of the culture. Similarly, the composition of the feed gas was measured daily, and the flow to the reactor was maintained at the required gas flow rate by using mass flow controllers. Once the H2 conversion reached 32%, the medium flow to the reactor was started at 1 ml / min, and 0.95 ml / min of permeate was withdrawn. The syngas flow rate to the reactor was increased based on the H2 conversion of the culture. If the H2 conversion was 32% or higher, the gas flow rate was increased by 10%. The cell mass was allowed to increase over time, reaching a cell mass of 3 g / L within 72 hours after inoculation of the reactor. At this point, the culture produced over 18 g / L ethanol. A liquid chromatograph was used to measure the ethanol and acetic acid concentrations in the broth. A gas chromatograph was used to measure the constituents of the syngas. The pH of the culture was maintained at about 4.5 using a neutralizing agent such as NH4OH. The cell density of the reactor was maintained at about 3 g / L by adjusting the permeate withdrawal rate. The reactor was set such that the permeate withdrawal rate inversely controlled the purge rate of cells from the reactor. The operating pressure was atmospheric. As the bacterial fermentation proceeds for several hours after inoculation, CO2 is produced from the conversion of CO and H2 is consumed along with CO. The production process and bacterial fermentation reactor system are then maintained at steady state with only minor adjustments from time to time to maintain acetate concentrations in the ranges described above and a cell density of approximately 3 g / L, producing 15-35 g / L ethanol and 0-5 g / L acetate as products. This continuous fermentation process allows for the continuous production and maintenance of high concentrations of ethanol with low concentrations of acetate by-product under stable operating conditions to enhance the use of the targeted bacteria on an industrial scale for ethanol production.

[0043] Example 8 Effect of acetic acid on Clostridium ljungdahlii Fermentations using Clostridium ljungdahlii were carried out as described in Example 7. Each test was carried out for 72 hours in the presence of 5 g / L NaCl and the following amounts of acetic acid in the reactor broth: Control - no addition Test 1 - 2g / L acetic acid Test 2 - 4g / L Acetic Acid Test 3 - 6g / L Acetic Acid The addition of acetate to the medium affected the specific productivity of ethanol. Thus, as the concentration of acetate increased, so did the amount of ethanol produced. There was no discernible change in acetate productivity, regardless of the concentration in the medium. All exogenously added acid appeared to be converted to ethanol by the cultures. When compared to the control, there was a significant improvement in the specific CO uptake in the cultures with exogenously added acid in the medium, with the highest concentration reaching the highest uptake level at about 0.750 mmol / min / g cells. The control averaged about 0.500 mmol / min / g cells during this time. Acetate concentration also increased hydrogen uptake. The specific productivity at steady state (g / L / day / g cells) was as follows:

[0044] [Table 4] Figure 12 illustrates the specific gas uptake by Clostridium ljungdahlii with various concentrations of acetate.

[0045] Example 9 Two-stage reactor using Clostridium ljungdahlii and Acetobacterium woodii The two-stage reactors were configured as shown in Figure 1. The two-stage reactors included stirred tank bioreactor Bi 120 and stirred tank bioreactor Bx 110. Each reactor contained liquid media containing vitamins, trace metals, cysteine ​​(as a sulfur source) and salts as described herein. Bioreactor Bi 120 started with actively growing Clostridium ljungdahlii and bioreactor Bx 110 started with actively growing Acetobacterium woodii. In bioreactor Bx 110, 0.5 M NaOH is used as the agent to maintain the pH at about 6.0. The estimated amount of NaOH used per gram of cells per hour is 0.21. Gaseous substrate Gi 186 containing 55% H2, 12% N2, 33% CO was introduced into bioreactor Bi 120. The outlet gas stream (Gp) from bioreactor Bi 120 was fed to bioreactor Bx 110 via gas flow connection 184. Water in the system exists in a "closed" configuration and passes from bioreactor 120 to distillation 188 via permeate / ethanol conduit 192. Water (distillation bottoms) is returned to bioreactor Bx 110 via water recycle line 202. Acetic acid from bioreactor Bx 110 is pumped to bioreactor Bi 120 via permeate acetate line 182. Fermentation was carried out for approximately 120 hours starting from water loop closure. The system was equilibrated for the first 58 hours and data was collected for the final 62 hours. The inlet gas streams 186 (Gi) and 187 (Gx) of both reactors and the outlet gas streams 184 (Gp) and 207 (exhaust gas stream of bioreactor Bx 110) were analyzed approximately every 4 hours. Analysis was performed using gas chromatography. Data was expressed as % composition. Analysis of ethanol, acetic acid and butanol products was performed approximately every 4 hours using liquid chromatography (LC). Cell concentration of both reactors was monitored approximately every 4 hours.

[0046] The average % composition and gas flow rates were as follows: [Table 5] The feed rates of the gas influent to bioreactors Bi 120 and Bx 110 were maintained using mass flow controllers (MFCs). The exit gas flow rates of bioreactors Bx 110 and Bi 120 were measured using a burette. The gas flow rate to bioreactor Bx 110 was calculated using the ratio of N2 composition of the influent and exhaust gas streams of bioreactor Bx 110. In this experiment, only a fraction of the gas exiting bioreactor Bi 120 was fed to bioreactor Bx 110. In one example, 51% of the effluent gas from bioreactor Bi 120 was fed to bioreactor Bx 110. In another example, 60% of the effluent gas from bioreactor Bi 120 was fed to bioreactor Bx 110. The ethanol productivity was as follows:

[0047] [Table 6] The specific ethanol productivity values ​​for the two reactor systems were calculated using the productivity of bioreactor Bi 120. Acetobacterium woodii produced zero to negligible amounts of ethanol in these experiments. The specific ethanol productivity of Clostridium ljungdahlii used in this table (without combining with bioreactor 110) was measured before the two reactors were coupled. If 100% of the off-gas from bioreactor Bi 120 was fed to bioreactor Bx 110, the specific ethanol productivity would be estimated to be 24.4 g / L / g Clostridium cells / day.

[0048] The average carbon capture rate of this system was: [Table 7] CO2 is produced in the bioreactor 120 via the water gas shift reaction. Figure 13 shows the utilization of CO1207 and CO21206 in bioreactor 110 when connected to bioreactor 120. Conversion is calculated using the molar concentration % of outgoing gas / incoming gas.

[0049] Example 10 Two-stage reactor using Clostridium ljungdahlii and Acetobacterium The two-stage reactor was configured as shown in FIG. 1. The two-stage reactor included stirred tank bioreactor 120 (Bi) and stirred tank bioreactor 110 (Bx). Each reactor contained liquid medium containing vitamins, trace metals, cysteine ​​(as a sulfur source) and salts as described herein. Bioreactor 120 (Bi) started with actively growing Clostridium ljungdahlii and bioreactor 110 (Bx) started with actively growing Acetobacterium woodii. In bioreactor Bx 110, 0.5 M NaOH is used as the agent to maintain the pH at about 5.55. The estimated amount of NaOH used per gram of cells per hour was 0.13 ml / min. Gaseous substrate 186 (Gi) containing 56% H2, 11% CH4, 33% CO was introduced into bioreactor Bi 120. The outlet gas stream (Gp) from bioreactor 120 was fed via gas flow connection 184 to bioreactor Bx 110. Water in the system is in a "closed" configuration and passes from bioreactor Bi 120 to distillation 188 via permeate / ethanol conduit 192. Water (distillation bottoms) is returned to bioreactor Bx 110 via water recycle line 202. Acetic acid from bioreactor Bx 110 is pumped to bioreactor Bi 120 via permeate acetate line 182. Fermentation was carried out for approximately 96 hours starting with the water loop closure. The system was equilibrated for the first 48 hours and data was collected for the final 48 hours. The inlet gas streams 186 (Gi) and 187 (Gx) and the outlet gas streams 184 (Gp) and 207 (vent streams) of both reactors were analyzed approximately every 4 hours. Analysis was performed using gas chromatography. Data was expressed as % composition. Analysis of the ethanol, acetic acid and butanol products was performed using GC approximately every 4 hours. The cell concentration of both reactors was monitored approximately every 4 hours.

[0050] The average % composition and gas flow rates were as follows: [Table 8] The feed rates of the gas influent to bioreactors Bi 120 and Bx 110 were maintained using mass flow controllers (MFCs). The exit gas flow rates of bioreactors Bx 110 and Bi 120 were measured using a burette. The gas flow rate to bioreactor Bx 110 was calculated using the CH4 composition ratio of the inlet and outlet gas streams of bioreactor Bx 110. In this experiment, only a fraction of the gas exiting bioreactor Bi 120 was fed to bioreactor Bx 110. The gas feed to bioreactor Bx 110 was 22.5% of the total exhaust gas from bioreactor Bi 120.

[0051] The ethanol productivity was as follows: [Table 9] The specific ethanol productivity values ​​for the two reactor system were calculated using the productivity of bioreactor Bi 120. Acetobacterium woodii produced zero to negligible amounts of ethanol in these experiments. The specific productivity by Clostridium ljungdahlii shown in this table (without combining with bioreactor Bx 110) was measured before the two reactors were coupled. If 100% of the off-gas from bioreactor Bi 120 was fed to bioreactor Bx 110, the specific ethanol productivity of this system is estimated to be 36.4 g / L / g Clostridium cells / day.

[0052] The butanol productivities were as follows: [Table 10] Butyric acid is produced by Acetobacterium woodii. This experiment shows that Clostridium ljungdahlii has the ability to convert butyric acid to butanol. If 100% of the exhaust gas from bioreactor Bi 120 is fed to bioreactor Bx 110, the specific productivity of butanol is estimated to be 8.2 g / L / g Clostridium cells / day.

[0053] The average carbon capture rate of this system was: [Table 11] CO2 is produced in the bioreactor Bi 120 by the water gas shift reaction.

[0054] While the disclosure disclosed herein has been described with reference to specific embodiments, examples and applications thereof, numerous modifications and variations may be made thereto by those skilled in the art without departing from the scope of the disclosure as set forth in the claims.

Claims

1. Supplying a gaseous substrate Gx to a bioreactor Bx, the gaseous substrate Gx being CO 2 and 5 to 90 mol % CO 2 The step of feeding acetogenic bacteria Mx to a bioreactor Bx, the acetogenic bacteria Mx comprising a sodium-transporting ATPase that is active during fermentation in the bioreactor Bx; Supplying sodium ions to the bioreactor Bx by one or more sodium ion sources; fermenting a gaseous substrate Gx with acetogenic bacteria Mx in a fermentation broth comprising acetogenic bacteria Mx and one or more sources of sodium ions to produce one or more organic acids, the fermentation broth comprising less than 0.01 grams of yeast extract per liter, less than 0.01 grams of carbohydrates per liter, sodium ions being supplied at a sodium supply rate of 290-8750 μg / gram of cells / minute, and the fermentation broth being maintained at a pH in the range of 4-6.9; feeding at least a portion of said one or more organic acids to a bioreactor Bi; Supplying a gaseous substrate Gi to a bioreactor Bi, the gaseous substrate Gi comprising CO and containing 20-90 mol% CO; Feeding acetogenic bacteria Mi to a bioreactor Bi, the acetogenic bacteria Mi comprising a proton-translocating ATPase that is active during fermentation in the bioreactor Bi; and In a fermentation broth containing acetogenic bacteria Mi, a gaseous substrate Gi in a bioreactor Bi is fermented by the acetogenic bacteria Mi to produce a liquid stream containing one or more alcohols and CO 2 generating a gas flow Gp comprising Including, The method, wherein the organic acid comprises acetic acid, the alcohol comprises ethanol, and the acetogenic bacterium Mi is a Clostridium bacterium.

2. The method of claim 1 , wherein the gaseous substrate Gx comprises at least a portion of the gas stream Gp.

3. 2. The method of claim 1, wherein an additional stream comprising one or more organic acids is fed to the bioreactor Bi.

4. The gaseous substrate Gx is H 2 The method of claim 1 , comprising:

5. 2. The method of claim 1, wherein the gaseous substrate Gx and the gaseous substrate Gi are selected from the group consisting of industrial gases, fermenter gas streams and mixtures thereof.

6. 2. The method of claim 1, wherein the acetogenic bacterium Mx is selected from the group consisting of Acetogenium kivui, Acetanaerobium noterae, Acetobacterium woodii, Alkalibaculum bacchi CP11, Moorella thermoacetica, Moorella thermoautotrophica, Ruminococcus productus, and combinations thereof.

7. The method according to claim 6, wherein the acetogenic bacterium Mx is Acetobacterium woodii.

8. 2. The method of claim 1, wherein the source of sodium ions is provided by a compound selected from the group consisting of sodium chloride, sodium hydroxide, sodium phosphate, sodium sulfate, sodium nitrate, sodium bicarbonate, sodium bisulfate and mixtures thereof.

9. 2. The method of claim 1, wherein the organic acid is one or more C1-C10 organic acids including acetic acid.

10. 10. The method of claim 9, wherein the organic acid is acetic acid.

11. The acetogenic bacteria Mi is Clostridium aceticum, Clostridium acetobutylicum, Clostridium autoethanogenum, Clostridium carboxydivorans, Clostridium drakei, Clostridium ljungdahlii, Clostridium magnum, Clostridium pasteurianum, 2. The method of claim 1, wherein the bacterial strain selected from the group consisting of Clostridium pasteurianum, Clostridium ragsdalei, Clostridium scatologenes, Clostridium thermoaceticum, Clostridium ultunenense, Clostridium stick-landii, and mixtures thereof.

12. The method according to claim 11, wherein the acetogenic bacterium Mi is Clostridium ljungdahlii.

13. Supplying a gaseous substrate Gx to a bioreactor Bx, the gaseous substrate Gx being CO 2 and H 2 and 5 to 90 mol % CO 2 The step of feeding acetogenic bacteria Mx to a bioreactor Bx, the acetogenic bacteria Mx comprising a sodium-transporting ATPase that is active during fermentation in the bioreactor Bx; Supplying sodium ions to the bioreactor Bx by one or more sodium ion sources; fermenting a gaseous substrate Gx with acetogenic bacteria Mx in a fermentation broth comprising acetogenic bacteria Mx and one or more sources of sodium ions to produce one or more organic acids, the fermentation broth comprising less than 0.01 grams of yeast extract per liter, less than 0.01 grams of carbohydrates per liter, sodium ions being supplied at a sodium supply rate of 290-8750 μg / gram of cells / minute, and the fermentation broth being maintained at a pH in the range of 4-6.9; feeding at least a portion of said one or more organic acids to a bioreactor Bi; Supplying a gaseous substrate Gi to a bioreactor Bi, the gaseous substrate Gi comprising CO and containing 20-90 mol% CO; Feeding acetogenic bacteria Mi to a bioreactor Bi, the acetogenic bacteria Mi comprising a proton-translocating ATPase that is active during fermentation in the bioreactor Bi; and In a fermentation broth containing acetogenic bacteria Mi, a gaseous substrate Gi in a bioreactor Bi is fermented by the acetogenic bacteria Mi to produce a liquid stream containing one or more alcohols and CO 2 generating a gas flow Gp comprising Including, The method, wherein the organic acid comprises acetic acid, the alcohol comprises ethanol, and the acetogenic bacterium Mi is a Clostridium bacterium.

14. The method of claim 13, wherein the gaseous substrate Gx comprises at least a portion of the gas stream Gp.

15. 14. The method according to claim 13, wherein an additional stream comprising one or more organic acids is fed to the bioreactor Bi.

16. 14. The method according to claim 13, wherein the gaseous substrate Gx and the gaseous substrate Gi are selected from the group consisting of industrial gases, fermenter gas streams and mixtures thereof.

17. The method of claim 13, wherein the acetogenic bacteria Mx is selected from the group consisting of Acetogenium kibui, Acetoanaerobium notelae, Acetobacterium woodii, Alkalibaculum bacchii CP11, Moorella thermoacetica, Moorella thermoautotrophica, Ruminococcus productus, and combinations thereof.

18. The method according to claim 17, wherein the acetogenic bacterium Mx is Acetobacterium woodii.

19. 14. The method of claim 13, wherein the source of sodium ions is provided by a compound selected from the group consisting of sodium chloride, sodium hydroxide, sodium phosphate, sodium sulfate, sodium nitrate, sodium bicarbonate, sodium bisulfate and mixtures thereof.

20. 14. The method of claim 13, wherein the organic acid is one or more C1-C10 organic acids including acetic acid.

21. 21. The method of claim 20, wherein the organic acid is acetic acid.

22. The method according to claim 13, wherein the acetogenic bacterium Mi is selected from the group consisting of Clostridium aceticum, Clostridium acetobutylicum, Clostridium autoethanogenum, Clostridium carboxydivorans, Clostridium drageei, Clostridium ljungdahlii, Clostridium magnum, Clostridium pasteurianum, Clostridium ragsdalei, Clostridium scatologenes, Clostridium thermoaceticum, Clostridium wurtunenense, and mixtures thereof.

23. The method according to claim 22, wherein the acetogenic bacterium Mi is Clostridium ljungdahlii.

24. Supplying a gaseous substrate Gx to a bioreactor Bx, the gaseous substrate Gx being CO 2 and 5 to 90 mol % CO 2 The step of feeding acetogenic bacteria Mx to a bioreactor Bx, the acetogenic bacteria Mx comprising a sodium-transporting ATPase that is active during fermentation in the bioreactor Bx; Supplying sodium ions to the bioreactor Bx by one or more sodium ion sources; fermenting a gaseous substrate Gx with acetogenic bacteria Mx in a fermentation broth comprising acetogenic bacteria Mx and one or more sources of sodium ions to produce one or more organic acids, the fermentation broth comprising less than 0.01 grams of yeast extract per liter, less than 0.01 grams of carbohydrates per liter, sodium ions being supplied at a sodium supply rate of 290-8750 μg / gram of cells / minute, and the fermentation broth being maintained at a pH in the range of 4-6.9; providing at least a portion of the one or more organic acids to a bioreactor system Bi-s; supplying a gaseous substrate Gi to a bioreactor system Bi-s, the gaseous substrate Gi comprising CO and containing 20-90 mol% CO; Feeding acetogenic bacteria Mi to a bioreactor system Bi-s, the acetogenic bacteria Mi comprising a proton-translocating ATPase that is active during fermentation in the bioreactor system Bi-s; and In a fermentation broth containing acetogenic bacteria Mi, a gaseous substrate Gi in a bioreactor system Bi-s is fermented by the acetogenic bacteria Mi to produce a liquid stream containing one or more alcohols and CO 2 generating a gas flow Gp comprising Including, The method, wherein the organic acid comprises acetic acid, the alcohol comprises ethanol, and the acetogenic bacterium Mi is a Clostridium bacterium.

25. 25. The method of claim 24, wherein the bioreactor system Bi-s comprises two or more bioreactors Bi-n.

26. 25. The method of claim 24, wherein the gaseous substrate Gi is supplied to two or more bioreactors Bi-n to achieve a gas superficial velocity effective to foam no more than 10% of the culture volume per hour.

27. 25. The method of claim 24, wherein the gaseous substrate Gx comprises at least a portion of the gas stream Gp.

28. The gaseous substrate Gx is H 2 25. The method of claim 24, comprising:

29. 25. The method of claim 24, wherein the gaseous substrate Gx and the gaseous substrate Gi are selected from the group consisting of industrial gases, fermenter gas streams and mixtures thereof.

30. 25. The method of claim 24, wherein the acetogenic bacteria Mx is selected from the group consisting of Acetogenium kibui, Acetoanaerobium notelae, Acetobacterium woodii, Alkalibaculum bacchii CP11, Moorella thermoacetica, Moorella thermoautotrophica, Ruminococcus productus, and combinations thereof.

31. The method according to claim 30, wherein the acetogenic bacterium Mx is Acetobacterium woodii.

32. 25. The method of claim 24, wherein the source of sodium ions is provided by a compound selected from the group consisting of sodium chloride, sodium hydroxide, sodium phosphate, sodium sulfate, sodium nitrate, sodium bicarbonate, sodium bisulfate and mixtures thereof.

33. 25. The method of claim 24, wherein the organic acid is one or more C1-C10 organic acids including acetic acid.

34. 34. The method of claim 33, wherein the organic acid is acetic acid.

35. 25. The method of claim 24, wherein the acetogenic bacteria Mi is selected from the group consisting of Clostridium aceticum, Clostridium acetobutylicum, Clostridium autoethanogenum, Clostridium carboxydivorans, Clostridium drageei, Clostridium ljungdahlii, Clostridium magnum, Clostridium pasteurianum, Clostridium ragsdalei, Clostridium scatologenes, Clostridium thermoaceticum, Clostridium wurtunenense, and mixtures thereof.

36. The method according to claim 35, wherein the acetogenic bacterium Mi is Clostridium ljungdahlii.

37. 25. The method of any one of claims 1, 13 or 24, wherein the organic acid is a combination of acetic acid and butyric acid and the alcohol is a combination of ethanol and butanol.

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