Co 2 upgrading into c 2 oxygenates with a cuag tandem electrocatalyst

Abstract

Electrochemical CO2 upgrading into C2 oxygenates is achieved a membrane electrode assembly (MEA) cell comprising metal plates, an anion exchange membrane (AEM), a cathode and anode, wherein the cathode comprises nanoscale Cu and Ag nanoparticles on a hydrophobic carbon substrate, wherein CO2 is fed to the cathodic side and converted to C2 oxygenates at the electrocatalyst surface.

Description

CO2 Upgrading into C2 oxygenates with a CuAg tandem electrocatalyst

[0001] Introduction

[0002] The chemical industry faces a significant challenge with CO2 emissions due to its heavy reliance on fossil fuels. Approaches involving electrochemical upgrading powered by renewable electricity have a potential to reduce these hard-to-abate emissions. Among CO2-derived products, C2 oxygenates products (ethanol and acetic acid) have a consistent global demand as they are feedstocks / precursors for fuel, value-added chemicals, and bioproducts. However, to date, CO2 upgrading to C2 oxygenates products at high efficiency remains elusive due to poor control over the reaction pathway.

[0003] In prior studies, we demonstrated that a tandem Cu-Ag electrocatalyst enhances the production rate of C2 oxygenate. Ag supplies CO intermediates to catalytic active sites, facilitating C-C coupling and shifting selectivity toward oxygenate products. Yet, these systems have been limited by low selectivity and reaction rates below the industrially relevant current density of > 100 mAcm-2.

[0004] Summary of the Invention

[0005] In an aspect the invention provides a tandem electrocatalyst and microenvironment within a membrane electrode assembly (MEA) system to investigate how CO generation / consumption switches the reaction pathway among C2 products. The MEA system, equipped with a gas diffusion electrode (GDE), directly supplies gas-phase CO2 to the electrocatalysts, allowing CO2 electrolysis to operate at a higher current density, approaching sub-A cm-2. Furthermore, reactant concentration can be tuned by exploiting mass transport control in the MEA system. This high production rate of C2 oxygenates enables collection of a concentrated liquid steam from the CO2 electrolyzer. The device can be used for CO2-to-C2 oxygenates for fuel synthesis, or bioproducts synthesis in combination with biological reactions.

[0006] In aspects the membrane electrode assembly (MEA) cell is employed with an anion exchange membrane (AEM). In the AEM / MEA system, liquid products cross the AEM: acetate, a negatively charged ion, is transported to the anodic side, while ethanol, a neutral species, is carried by electro-osmotic force. Thus, concentrated C2 oxygenates liquid can be achieved in the anolyte, allowing for being a feedstock / substrate for longer-chain carbon molecules.

[0007] In aspects the invention provides a tandem CuAg electrocatalyst and leverages the microenvironment in the MEA system to achieve CO2 upgrading into C2 oxygenates with a high reaction rate. Cu is the material that produces C2products in the CO2electrolysis system. While Cu is well-known to produce C2 products, it typically favors ethylene over oxygenates. For C21 B29-099-2WOoxygenates production, the catalyst design is essential to shift the product distribution toward C2 oxygenates. By embedding Ag nanodomain within Cu, they form a close network. In this network, local CO generation from Ag enhances *CO coverage on Cu, shifting the reaction pathway toward C2 oxygenates production. We then control CO / CO2 mass transport at the catalyst surface by using the gas-diffusion electrode and tunable CO2flux. This enables precise management *CO residence time at the active sites and the creation of the CO-enriched microenvironment. This combined catalyst and microenvironment design favors acetate and ethanol generation at a high current density of > 100 mA cm-2.

[0008] Additionally, the MEA system, paired with AEM, selectively transports and concentrates C2 oxygenates into the anolyte. Under industrially relevant conditions, the CO2 electrolysis system yields a highly concentration of C2 oxygenates that can serve as a feedstock for subsequent biological reactions.

[0009] Our integrated catalyst and microenvironment design enables us to electrosynthesize C2 oxygenates at industrially relevant production rates. We achieved 44% C2 oxygenates FE, with 26% for CH3COO‘ and 18% for C2H5OH, at 400 mA cm-2and -3.9 Vfull-cell. The electrolysis system continuously generates the acetate-rich stream in the bio-compatible electrolyte, which can be subsequently fed to the bioreactor. The invention provides optimizing catalyst stability and selectivity, scaling up the reactor design, and coupling this process with biological pathways for sustainable chemical production.

[0010] In aspects and embodiments the invention provides:[Oil] A device for electrochemical CO2 upgrading into C2 oxygenates, comprising a membrane electrode assembly (MEA) cell, the cell comprising metal plates, an anion exchange membrane (AEM), a cathode and anode, wherein the cathode comprises nanoscale Cu and Ag nanoparticles on a hydrophobic carbon substrate, wherein CO2 is fed to the cathodic side and converted to C2 oxygenates at the electrocatalyst surface, substantially as disclosed herein, e.g. Fig. 1A, 3C, 4A, 4B.

[0012] A method for electrochemical CO2 upgrading into C2 oxygenates, comprising operating a device herein.

[0013] A device or method herein configured to produce CO2-to-C2 oxygenates for fuel synthesis, or bioproducts synthesis in combination with biological reactions.

[0014] A device or method herein configured with formulations herein, or with CO2 to C2 efficacies wherein, especially for liquid C2 products.

[0015] A method for upgrading CO2 into a biopolymer comprising using a two-step abiotic-biotic system, comprising an electrolysis system, wherein CO2 is converted into C2 oxygenates (such as acetate and ethanol) at the cathode, using a Cu-Ag tandem electrocatalyst, the2 B29-099-2WOelectrocatalyst comprising a membrane electrode assembly (MEA) cell, the cell comprising metal plates, an anion exchange membrane (AEM), a cathode and anode, wherein the cathode comprises nanoscale Cu and Ag nanoparticles on a hydrophobic carbon substrate, wherein CO2 is fed to the cathodic side and converted to C2 oxygenates at the electrocatalyst surface, wherein the electrolysis process generates a liquid stream containing oxygenates in a bio-compatible electrolyte (anolyte), wherein these electrosynthesized oxygenates are then fed to a bioreactor, where substrates are upgraded to the biopolymer.

[0016] A method herein, wherein the biopolymer is 3-polyhydroxybutyrate (PHB).

[0017] A method herein, wherein the biopolymer is 3-polyhydroxybutyrate (PHB), wherein PHB is biosynthesized from acetate in a Cupriavidus necator (C. nectar) bioreactor, wherein acetate is taken up by C. necator and converted to acetyl-CoA via acetyl-CoA synthetase (acs), wherein acetyl-CoA enters the TCA cycle for energy generation or serves as a precursor for PHB biosynthesis, wherein in the PHB biosynthetic pathway, PhaA catalyzes the condensation of two molecules of acetyl-CoA to form acetoacetyl-CoA, which is then reduced to 3-hydroxybutyryl-CoA by PhaB. Finally, PhaC polymerizes 3-hydroxybutyryl-CoA into polyhydroxybutyrate (PHB).

[0018] A method herein, wherein the C. nectar is engineered to express genes encoding enzymes involved in PHB biosynthesis.

[0019] A method herein, wherein the system is configured substantially as disclosed in Fig. 1 A, 3C, 4A and / or 4B.

[0020] A method herein, wherein the system is configured to produce CO2-to-C2 oxygenates for fuel synthesis, or bioproducts synthesis in combination with biological reactions.

[0021] A method herein, wherein the system is configured with formulations herein, or with CO2 to C2 efficacies wherein, especially for liquid C2 products.

[0022] The invention encompasses all combinations of the particular embodiments recited herein, as if each combination had been laboriously recited.

[0023] Brief Description of the Drawings

[0024] Fig 1A-C. CO2 upgrading into bioproducts using the abiotic-biotic system, (a) Process flow for abiotic-biotic conversion of CO2 to chemicals and fuels. CO2 is electrochemically upgraded into C2oxygenates (such as acetate and ethanol) at the cathode, which are then concentrated in a bio-compatible anolyte. These oxygenates are supplied to a bioreactor where bacteria convert these substrates into valuable bioproducts, (b) Schematic of PHB biosynthesis from acetate in Cupriavidus necator (C. nectar). Acetate is taken up by C. necator and converted to acetyl-CoA via acetyl-CoA synthetase (acs). Acetyl-CoA enters the 3 B29-099-2WOTCA cycle for energy generation or serves as a precursor for PHB biosynthesis. In the PHB biosynthetic pathway, PhaA catalyzes the condensation of two molecules of acetyl-CoA to form acetoacetyl-CoA, which is then reduced to 3-hydroxybutyryl-CoA by PhaB. Finally, PhaC polymerizes 3 -hydroxybutyryl- Co A into polyhydroxybutyrate (PHB). Blue text indicates genes encoding enzymes involved in PHB biosynthesis.36(c) Carbon footprint comparison between petroleum-based polymers and the two-step system. The two-step system involves the abiotic-biotic conversion of CO2 into polyhydroxybutyrate (PHB), with energy efficiency (EE) referring to the CO2 electrolyzer. In both cases, the yield of the bioreactor is assumed to be 50%.

[0025] Fig 2A-F. Morphology and characterization of the Cu-Ag tandem electrocatalyst, (a-b) Transmission electron microscopy images of (a) 7 nm Cu nanoparticles and (b) 6 nm Ag nanoparticles, (c-d) Scanning electron microscope images of (c) as-prepared Cu-Ag tandem electrocatalyst on carbon paper-based gas diffusion electrode and (d) post-electrolysis of Cu-Ag tandem electrocatalyst on carbon paper-based gas diffusion electrode, (e-f) X-ray photoelectron spectroscopy spectra of (e) Cu of as-prepared Cu-Ag tandem electrocatalyst and (f) Ag of as-prepared Cu-Ag tandem catalyst.

[0026] Fig 3A-C. CO coverage-dependent selectivity of C2 products, (a-b) product generation rate and the ratio between C2 oxygenates and C2H4 for different CO2 flux at applied current density 300 mA cm’2in the membrane electrode assembly cell (a) with pure Cu electrocatalyst (b) with Cu-Ag (Ag 32%) tandem electrocatalyst, (c) Schematic of species flow and CO coverage on the electrocatalyst surface under CO2-rich and CO2-lean conditions. A CO2-rich condition corresponds to high CO2 flux, while a CO2-lean condition corresponds to low CO2 flux.

[0027] Fig 4A-B. a) Schematic of a membrane electrode assembly (MEA) cell. The cell components are metal plates, an anion exchange membrane (AEM), a cathode and an anode, b) Liquid product crossover in the anion exchange membrane (AEM) / MEA system. Formate and acetate are transported to the anode as they are negatively charged ions, and neutral molecules, such as ethanol and propanol, are dragged by electro-osmotic force.

[0028] Description of Particular Embodiments of the Invention

[0029] Unless contraindicated or noted otherwise, in these descriptions and throughout this specification, the terms “a” and “an” mean one or more, the term “or” means and / or. It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and4 B29-099-2WOscope of the appended claims. All publications, patents, and patent applications cited herein, including citations therein, are hereby incorporated by reference in their entirety for all purposes.

[0030] Catalyst preparation

[0031] 7 nm Cu and 6 nm Ag nanoparticles were synthesized using our group' s published methods. Cu nanoparticles were synthesized through colloidal synthesis. 0.5 mmol of copper(I) acetate and 0.5 mmol of tetradecylphosphonic acid (TDPA) were added to trioctylamine (TOA) solvent (10ml) at room temperature. TOA solvent was pre-heated at 130°C for 30 minutes under a nitrogen atmosphere to remove moisture. While stirring and keeping it under a nitrogen atmosphere, the solution was heated to 180°C for 30 min and then to 270°C for 30 min.Subsequently, the solution was cooled until room temperature. Ethanol was added to separate nanoparticles from the solvent, and the solution mixture was centrifuged at 6000 rpm for 15 minutes. Separated nanoparticles were washed with chloroform and acetone and redispersed in hexane. Ag nanoparticles were synthesized with a similar method. 10 mL of trioctylamine was purged with nitrogen gas at 130 °C for 30 min and cooled to room temperature, after which 0.50 mmol of silver(I) acetate and 0.25 mmol of TDPA were added. The solvent was heated with stirring to 130 °C for 1 h under a N2 atmosphere. After the reaction, the heating mantle was removed, and the solution was cooled to room temperature. Ethanol was added to separate nanoparticles from the solvent, and the solution mixture was centrifuged at 6000 rpm for 15 minutes. Ag nanoparticles were redispersed in hexane and acetone was added dropwise until the solution became turbid for post-size selection. After centrifugation at 12,000 r.p.m. for 10 min, Ag NPs were redispersed in hexane.

[0032] The concentration of Cu and Ag was analyzed by inductively coupled plasma optical emission spectroscopy (PerkinElmer Optima 7000 DV). The mixed solution with different Cu-Ag ratios was prepared considering the concentration of Cu and Ag.

[0033] The sizes and shapes of the as-prepared NPs were confirmed by transmission electron microscopy (Hitachi H-7650) and scanning electron microscopy (Ultra 55-FESEM). The surface composition was measured by X-ray photoelectron spectroscopy with an Al Kα source using a Thermo Scientific K- Alpha instrument.

[0034] Electrode preparation

[0035] The cathode was prepared by airbrushing graphite ink and Cu-Ag ink onto carbon paper (GDS5130, AvCarb). 20 um graphite was dispersed with Nafion binder in Isopropyl alcohol (1PA), and then the ink was sonicated for 1 hour. The mass loading of graphite is 0.05 mg cm’2. The mixed Cu-Ag NPs were dispersed in Hexane, and the solution was sprayed onto the graphite / carbon paper until the mass loading reached 0.1 mg cm’2. The cathode was dried for 24 hours in a vacuum chamber.5 B29-099-2WO

[0036] Electrochemical experiments

[0037] The electrochemical data were collected using an electrochemical station (biologies). The 5 cm-2membrane electrode assembly cell (dioxide materials) was used. The cathode and anode were separated by an anion exchange membrane (Sustainion® X37-50 grade RT Membrane). The anode, an oxygen evolution reaction catalyst, is titanium felt- supported iridium oxide (Magneto). The anolyte is 0.1 M CSH2PO4 or KH2PO4 and it was circulated using a peristaltic pump. CsH2PO4 was prepared by mixing Cs2CO3 or CsHCO3 and H3PO4. Humidified CO2 gas is fed with a mass flow from 2 to 10 sccm / cm2. Electrolysis was maintained for at least 30 min before collecting gas and liquid samples. The experiments were repeated at least 3 times for the average and standard error.

[0038] The gas-phase products were analyzed by gas chromatography (Agilent Technologies, 7890B) equipped with a thermal conductivity detector (TCD) and a flame ionization detector (FID). The liquid phase products were analyzed with a 500 MHz Bruker Avance IV NEO.

[0039] Results in brief

[0040] We prepared nanoscale copper and silver nanoparticles (NPs) and employed the Cu-Ag bimetallic NPs on the carbon paper-based GDE. The 7 nm Cu NPs and 6 nm Ag NPs were synthesized via colloidal synthesis, with a narrow size distribution, as shown in Transmission electron microscopy (TEM) images (Fig. 2a, b). The as-synthesized Cu and Ag NPs were mixed in hexane and coated on the GDE using an air-brush method. Scanning electron microscopy (SEM) images show that the Cu-Ag NPs assembly was uniformly distributed on the carbon paper (Fig. 2c), and Cu-Ag NPs forms a close-contact network during electrolysis (Fig. 2d). In previous work, our group found that Cu and Ag NPs migrate and agglomerate, as revealed by operando and post-electrolysis microscopic studies. This results in Ag nanodomains becoming embedded within the Cu domains. In this network, Ag serves as a CO-producing electrocatalyst, while C-C coupling occurs at metallic nanodomains, producing a mixture of C2+ products (acetate, ethylene, ethanol and propanol). To investigate how CO generation / consumption rate affects the selectivity in the CO2 reduction reaction (CO2RR) systems, we prepared various ratios of Ag at 0%, 19%, 32% and 58%. The surface composition of Cu-Ag was confirmed by X-ray photoelectron spectroscopy (XPS) (Fig. 2e, f).

[0041] We conducted CO2 electrolysis using Cu-Ag tandem electrocatalysts in the MEA system. The GDE with Cu-Ag tandem electrocatalysts, the AEM and IrOx / Ti felt were pressurized between two metal plates. High-purity CO2 gas was fed to the cathode, and 0.1 M CSH2PO4 was used as the anolyte. We then applied constant current in the range of 100-400 mA cm'2to the MEA cell. However, CO was the dominant product while C2oxygenates FE remained < 20% when using Cu-Ag electrocatalysts at a CO2 flow rate of > 3.4 µmol s-1cm-1(56 B29-099-2WOsccm cm-2). We posited that the limited residence time of CO at the electrocatalyst surface, due to the high CO2 flux, prevented CO from being consumed for C-C coupling. We tested this hypothesis by decreasing the CO2 flow rate until all CO2 could be consumed for C2 product formation; for example, the minimum CO2flux for 8e’ products and 12e’ products is 2.1 pinol s’1cm'1at 300 mA cm’2, with a theoretical CO2 conversion efficiency at 25-36% in the alkaline CO2 electrolysis system. In the CO2 flux range of 1-7 pmol s’1cm’1, we observed an increase in C2 FE as CO2 flux decreased for both Cu and Cu-Ag systems (Fig. 3a, b). As CO2 flux diminishes, CO molecules remain longer at C-C coupling active sites, enhancing C2 conversion efficiency. Product distribution was varied with different electrocatalysts. For Cu, C2H4 is the major product, with FE of 34-49%, while C2oxygenates production overwhelms C2H4 by using Cu-Ag (Ag 32%). The ratio of C2 oxygenates and C2H4 (C2 oxygenates / C2H4) is higher for Cu-Ag than for Cu at all CO2 inputs. Among C2 oxygenates, a substantial increase in CH3COO’ production was observed, followed by C2H5OH, as the CO2 flux decreased from 6.7 pniol s’1cm’1to 1.8 pmol s’1cm’1. The highest FE for C2 oxygenates was achieved with CH3COO’ FE of 21% and C2H5OH FE of 23% at the CO2 flux of 1.8 pmol s’1cm’1. The shift in selectivity from C2H4 to CH3COO’ in the tandem system is driven by the increased rate of CO generation rate from Ag, which creates CO-enriched microenvironments and boosts *CO coverage on the Cu-Ag surface (Fig. 3c). In contrast, pure Cu catalysts lack internal CO production, leading to a constant C2 oxygenates / C2H4 across CO2 flux.

[0042] We further studied Cu-Ag ratios to gain insights into how *CO coverage tunes the reaction pathway of C2 products. Pure Cu and Cu-Ag electrocatalysts with Ag contents of 19%, Ag 32% and Ag 58% were tested at 1.8 pmol s’1cm’1and an applied current density of 300 mA cm’2. Increasing Ag content at the catalytic sites boosts the CO generation, which in turn raises the C2 oxygenates / C2H4 from 0.4 to 3.7. At higher Ag concentration (58%), total C2 FE drops due to a lack of C-C coupling sites; however, the C2oxygenates / C2H4is 3.7, compared to 0.4 for pure Cu. These results suggest that *CO coverage, enhanced by CO-producing catalysts, promotes C-C coupling, particularly towards CH3COO’ rather than C2H4. As a result, using the Cu-Ag (Ag 32%) tandem electrocatalyst under limited CO2 flux conditions, we achieved 44% C2 oxygenates FE, with 26% for CH3COO’ and 18% for C2H5OH, at 400 mA cm’2and -3.9 VfuU-Ceii. The partial current density for C2 oxygenate products reaches 176 mA cm’2.

[0043] We sought to collect C2 oxygenate products in the bio-compatible anolyte (0.1 M KH2PO4) until their concentration reached levels suitable for subsequent biological reactions. We collected 200 mM of acetate and 80 mM of ethanol in 30 mL anolyte over 4 hours at the applied current density of 300 mA cm’2, with the C2 oxygenates FE in the range of 35-39%. The concentration of C2 oxygenates steadily increases, with no significant loss of CH3COO’ or 7 B29-099-2WOC2H5OH due to oxidation. The pH of the anolyte decreases from 4.6 to 2.7, but the full-cell voltage remained stable, with only a deviation of ~0.3 V. We circulated the anolyte at a volume of 6 mL cm-2to achieve the concentration of C2 oxygenate -60 mM h’1and replaced it with fresh solution whenever the concentration of C2 oxygenates reached desired level of > 200 mM.

[0044] Example: CO2 Upgrading into Bioproducts Using a Two-Step Abiotic-Biotic System

[0045] The chemical industry faces a significant challenge with CO2 emissions due to its heavy reliance on fossil fuels. While many strategies for replacing fossil fuels with biomass feedstocks for biological fermentation have been proposed and are in various stages of development,1'3they often compete with the human food chain and require extensive land use.4’6Alternatively, an approach involving electrochemical upgrading powered by renewable electricity offers the potential to reduce hard-to-abate CO2 emissions. Yet, to date, direct electrosynthesis of long-chain molecules (C4+) from CO2 has only achieved a current density of < 2 mA cm'2,7'10compared to C2 oxygenates (acetate and ethanol) that have reached industrially relevant current densities exceeding 100 mA cm-2.11-13Such high current densities motivated us to explore an approach combining an electrochemical system for CO2-to-C2 oxygenate conversion with a microbial system that utilizes C2 oxygenates to produce chemicals beyond C1-2 products. The electrosynthesized C2 oxygenates serve as viable substrates for microorganisms, allowing them to consume a liquid feedstock more efficiently than gaseous molecules and produce diverse value-added products.14’16

[0046] Several studies have explored CO2 conversion to bioproducts using a sequential process of CO2 electrolysis followed by microbial fermentation. ’ " Among CO2-de rived products, acetate has been considered a key substrate since it forms acetyl-CoA that participates in various biological valorization processes.14’23 25The acetate stream is typically generated through a tandem process: electrolysis of CO2 to CO, followed by CO-to-acetate conversion, due to the unfavorable reaction pathway of CO2 to acetate.26'29However, this approach adds complexity to the system and increases the costs associated with CO / CO2 separation.24, 30, 31Furthermore, the downstream operation of the CO2 electrolyzer poses a challenge, as liquid products must be separated from a concentrated electrolyte (-10 M), which is detrimental to microorganisms due to excessive salts.32Previous studies addressed this issue by introducing the adjustable effluent in an additional layer or using a diluted electrolyte.20’22However, these strategies increase resistance within the systems, resulting in higher costs.24Therefore, selecting a bio-comparable electrolyte while maximizing the overall system efficiency is a key design principle for coupling CO2electrolysis with microbial fermentation.8 B29-099-2WO

[0047] Here, we present a proof-of-concept demonstration of a two-step abiotic-biotic system that upgrades CO2 into 3-polyhydroxybutyrate (PHB) (Fig. la). First, CO2 electrolysis delivers sufficient C2 oxygenates under industrially relevant operating conditions at > 100 mA cm'2 3"'3’ To this end, we employed a Cu-Ag tandem electrocatalyst and a membrane electrode assembly (MEA) cell with an anion exchange membrane (AEM). In the AEM / MEA system, liquid products cross the AEM; acetate, a negatively charged ion, is transported to the anodic side, while ethanol, a neutral species, is carried by electro-osmotic force. (Fig. 4) The generated liquid stream from the anodic outlet, a mixture of C2 oxygenates and anolyte, is a bio-compatible solution. In the subsequent bio-upgrading stage, Cupriavidus necator is employed to utilize the acetate in the electrosynthesized solution as a carbon source for PHB production. In this process, C. necator converts acetate to acetyl-CoA by acetyl-coA synthetase, and the resulting acetyl-CoA enters C. necator s native metabolic pathway involving three enzymes encoded in one operon, PhaA, PhaB, and PhaC, to synthesize PHB (Fig. lb).36The abiotic-biotic system, which divides the process into two separate steps (converting CO2 to acetate, then converting acetate to PHB) circumvents challenges such as media conditions unsuitable for bacterial growth, thus allowing each step to be optimized independently.

[0048] The abiotic-biotic system couples electrochemical CO2 conversion with biological polymer synthesis, offering a promising route for sustainable biopolymer production. Herein we offer an analysis that estimates the carbon footprint of the two-step system for biopolymer production (Fig. 1c). For large-scale commodity production, microbial productivity is projected to reach 2-4 g L' 1 h' 1.16 ’ 379Assuming a 50% yield in the biorcactor and a volumetric flow of 5 mL per cm2active area in the CO2 electrolyzer,37’39the required reaction rate of C2 oxygenates is > 72 mA cm', contributing to 0.7-2 kg CO2eq / kg polymer at a 20% energy efficiency of CO2 electrolyzer, depending on the renewable electricity sources used (Supplementary Note 1 and 2). The major source of carbon emission is the CO2 electrolyzer, as the bioreactor has relatively low heat and electricity demand. This carbon footprint is lower than that of petroleum-based polymers, which is ~4 kg CO2eq / kg polymer.40This estimate suggests that abiotic-biotic upgrading has the potential to produce carbon-neutral chemicals.

[0049] In prior studies, we demonstrated that a tandem Cu-Ag electrocatalyst enhances the production rate of C2 oxygenate.41, 42Ag supplies CO intermediates to catalytic active sites, facilitating C-C coupling and shifting selectivity toward oxygenate products. Yet, these systems have been limited by low selectivity and reaction rates below the target current density of > 72 mA cm-2(Supplementary Note 1). The MEA system, equipped with a gas diffusion electrode (GDE), directly supplies gas-phase CO2to the electrocatalysts, allowing CO2 electrolysis to9 B29-099-2WOoperate at a higher current density, approaching sub-A cm-2. Furthermore, reactant concentration can be tuned by exploiting mass transport control.43-45

[0050] We proceeded by implementing these concepts in combination. We prepared nanoscale copper and silver nanoparticles (NPs) using our group’s prior method46, 47and employed the Cu-Ag bimetallic NPs on the carbon paper-based GDE. The 7 nm Cu NPs and 6 nm Ag NPs were synthesized via colloidal synthesis, with a narrow size distribution, as shown in transmission electron microscopy (TEM) images (Fig. 2a, b). The as-synthesized Cu and Ag NPs were mixed in hexane and coated on the GDE using an air-brush method. Scanning electron microscopy (SEM) images show that the Cu-Ag NPs assembly was uniformly distributed on the carbon paper (Fig. 2c), and Cu-Ag NPs form a close-contact network during electrolysis (Fig. 2d). In previous work, our group found that Cu and Ag NPs migrate and agglomerate, as revealed by operando and post-electrolysis microscopic studies. This results in Ag nanodomains becoming embedded within the Cu domains.42In the network, Ag serves as a CO-producing electrocatalyst, while C-C coupling occurs at metallic nanodomains, producing a mixture of C2+ products (acetate, ethylene, ethanol, and propanol). To investigate how CO generation / consumption rate affects the selectivity in the CO2 reduction reaction (CO2RR) systems, we prepared various ratios of Ag at 0%, 19%, 32%, and 58%. The surface composition of Cu-Ag was confirmed by X-ray photoelectron spectroscopy (XPS) (Fig. 2e, f).

[0051] We conducted CO2 electrolysis using Cu-Ag tandem electrocatalysts in the MEA system. The GDE with Cu-Ag tandem electrocatalysts, the AEM, and IrOx / Ti felt were pressurized between two metal plates. High-purity CO2 gas was fed to the cathode, and 0.1 M CSH2PO4 was used as the anolyte. We then applied constant current in the range of 100-400 mA cm'2to the MEA cell. However, CO was the dominant product while C2 oxygenates EE remained < 20% when using Cu-Ag electrocatalysts at a CO2 flow rate of > 3.4 pmol s1cm'1(5 sccm cm-2). We posited that the limited residence time of CO at the electrocatalyst surface, due to the high CO2 flux, prevented CO from being consumed for C-C coupling.43’44We tested this hypothesis by decreasing the CO2 flow rate until all CO2 could be consumed for C2 product formation; for example, the minimum CO2flux for 8e' products and 12e' products is 2.1 pniol s'1cm'1at 300 mA cm'2with a theoretical CO2 conversion efficiency in the alkaline CO2 electrolysis system (Supplementary Note 3)48In the CO2 flux range of 1-7 µmol s-1cm-1, we observed an increase in C2 FE as CO2 flux decreased for both Cu and Cu-Ag systems (Fig. 3a, b). As CO2 flux diminishes, CO molecules remain longer at C-C coupling active sites, enhancing C2 conversion efficiency. However, H2 FE increases at the point where CO2 is fully depleted. Product distribution was varied with different electrocatalysts. For Cu, C2H4 is the major product, with FE of 34-49%, while C2 oxygenates production overwhelms C2H4 by using Cu-Ag 10 B29-099-2WO(Ag 32%). The ratio of C2 oxygenates and C2H4 (C2 oxygenates / C2H4) is higher for Cu-Ag than for Cu at all CO2 inputs. Among C2 oxygenates, a substantial increase in CH3COO" production was observed, followed by C2H5OH, as the CO2 flux decreased from 6.7 pmol s’1cm"1to 1.8 pmol s1cm"1. The highest FE for C2 oxygenates was achieved with CH3COO FE of 21% and C2H5OH FE of 23% at the CO2 flux of 1.8 pmol s'1cm’1. The shift in selectivity from C2H4 to CII3COO’ in the tandem system is driven by the increased rate of CO generation rate from Ag, which creates CO-enriched microenvironments and boosts *CO coverage on the Cu-Ag surface (Fig. 3c).45’49’50In contrast, pure Cu catalysts lack internal CO production, leading to a constant C2 oxygenates FE across CO2 flux (Fig 3a).

[0052] We further studied Cu-Ag ratios to gain insights into how *CO coverage tunes the reaction pathway of C2 products. Pure Cu and Cu-Ag electrocatalysts with Ag contents of 19%, Ag 32%, and Ag 58% were tested at 1.8 pmol s'1cm"1and an applied current density of 300 mA cm"2. Increasing Ag content at the catalytic sites boosts the CO generation, which in turn raises the C2 oxygenates / C2H4 from 0.4 to 3.7. At higher Ag concentration (58%), total C2 FE drops due to a lack of C-C coupling sites; however, the C2 oxygenates / C2H4 is 3.7, compared to 0.4 for pure Cu. These results suggest that *CO coverage, enhanced by CO-producing catalysts, promotes C-C coupling, particularly towards CH3COO’ rather than C2H4. As a result, using the Cu-Ag (Ag 32%) tandem electrocatalyst under limited CO2 flux conditions, we achieved 44% C2 oxygenates FE, with 26% for CH3COO’ and 18% for C2H5OH, at 400 mA cm"2and -3.9 VfuU-ceii- The partial current density for C2 oxygenate products reaches 176 mA cm"2, exceeding the target production rate of 72 mA cm’2for microbial biotic upgrading on an industrial scale (Supplementary Note 1).

[0053] We sought to collect C2 oxygenate products in the bio-compatible anolyte (0.1 M KH2PO4) until their concentration reached levels suitable for microorganisms. We collected 200 mM of acetate and 80 mM of ethanol in 30 mL anolyte over 4 hours at the applied cunent density of 300 mA cm'2, with the C2 oxygenates FE in the range of 35-39%. The concentration of C2 oxygenates steadily increases, with no significant loss of CH3COO’ or C2H5OH due to oxidation. The pH of the anolyte decreased from 4.6 to 2.7, but the full-cell voltage remained stable, with only a deviation of ~0.3 V. We circulated the anolyte at a volume of 6 mL cm2to achieve the concentration of C2 oxygenate -60 mM h"1and replaced it with fresh solution whenever the concentration of C2 oxygenates reached desired level of > 200 mM. In the solution, inorganic species, such as Cu, Ag and Ir, were detected at concentrations below -100 ppb (Table S2).

[0054] As a proof-of-concept demonstration of a two-step system, we sought to feed electrosynthesized acetate to C. necator to produce PHB on a laboratory scale. The results of the 11 B29-099-2WObioconversion process demonstrate PHB production by 24 h, when the PHB concentration reached 433.97 ± 31.16 mg E1and the cell concentration increased to 0.141 ± 0.0065 g Ll. The corresponding PHB content of the cells was 75.4 ± 1.6 weight %. The highest PHB production rate was observed between 12 and 22 h, reaching 32.35 ± 3.49 mg I.1h To further investigate intracellular PHB accumulation, confocal fluorescence microscopy was performed after staining the cells with Nile red, a lipophilic dye that selectively binds to PHB granules. The confocal images clearly show distinct fluorescence signals within the cells, qualitatively confirming PHB formation. This visual evidence supports the quantitative measurements, demonstrating that the electrosynthesized acetate solution effectively sustains microbial PHB biosynthesis.Furthermore, to obtain PHB in purified form, the polymer was extracted from harvested cells using 1,3 -dioxolane, a green solvent known for its efficient PHB dissolution.51The extraction yielded 61 mg of PHB as a fine powder, further demonstrating its successful biosynthesis and accumulation. These findings highlight the power of the two-step abiotic-biotic system for sustainable and carbon-neutral chemical production, especially for converting CO2 into value-added biopolymers

[0055] The combination of catalyst design and microenvironment control enables electrosynthesis of C2 oxygenates at industrially relevant production rates. The electrolysis system continuously generates the acetate-rich stream in the bio-compatible electrolyte, which may be subsequently fed to C. necator in the bioreactor.

[0056] Electrochemical experiments

[0057] The electrochemical data were collected using an electrochemical station (biologies). The 5 cm’2membrane electrode assembly cell (dioxide materials) was used. The cathode and anode were separated by an anion exchange membrane (Sustainion® X37-50 grade RT Membrane). The anode, an oxygen evolution reaction catalyst, is titanium felt- supported iridium oxide (Magneto). The anolyte is 0.1 M CsH2PO4 or KH2PO4. and it was circulated using a peristaltic pump. CsH2PO4 was prepared by mixing Cs2CO3 or CsHCO3 and H3PO4. Humidified CO2 gas is fed with a mass flow from 2 to 10 sccm / cm2. Electrolysis was maintained for at least 30 min before collecting gas and liquid samples. The experiments were repeated at least 3 times for the average and standard error.

[0058] The gas-phase products were analyzed by gas chromatography (Agilent Technologies, 7890B) equipped with a thermal conductivity detector (TCD) and a flame ionization detector (FID). 1’he liquid phase products were analyzed with a 500 MHz Broker Avance IV NEO.

[0059] Microbial experiments

[0060] Cupriavidus necator (ATCC 17699) was obtained from the American Type Culture Collection (ATCC). Prior to experimentation, a single colony was inoculated into 5 mL of 12 B29-099-2WOLuria-Bertani (LB) broth and cultivated overnight at 30°C, 200 rpm. The overnight culture was used to inoculate 50 mL of M9 minimal medium containing fructose as the carbon source in a 250-mL baffled Erlenmeyer flask. The culture was incubated at 30°C, 200 rpm for 12-16 hours until the exponential growth phase was reached. Cells were harvested by centrifugation at 4,000 x g for 10 min at 4°C, washed twice with sterile phosphate-buffered saline (PBS, pH 7.0), and resuspended in fresh M9 containing electrochemically synthesized acetate (3 g L1) as the sole carbon source.52The initial optical density at 600 nm (OD6oo) was adjusted to 0.5. Nitrogen limitation was applied to promote polyhydroxybutyrate (PHB) accumulation by supplementing ammonium chloride (NH4CI) at a concentration of 0.09 g L1, maintaining a carbon-to-nitrogen (C / N) molar ratio of approximately 30.5All cultivations were performed in triplicate. Bacterial growth and PHB production were monitored at designated time points by measuring OD600and PHB concentrations.

[0061] Product analysis using high-performance liquid chromatography (HPLC)

[0062] Cell pellets were collected from 5 mL of culture by centrifugation at 4,000 x g for 10 min. The harvested pellets were lysed and the PHB was depolymerized by adding 1 mL of concentrated sulfuric acid (H2SO4) and incubating at 95°C for 60 min. The depolymerized PHB solution was then diluted with 4 mL of HPLC-grade water and following the addition of adipic acid solution as an internal standard filtered through a 0.2 pm filter prior to analysis. PHB was quantified using a high-performance liquid chromatography system (HPLC 1260 Infinity II, Agilent, CA) equipped with an Aminex HPX-87H column (300 x 7.8 mm; Bio-Rad). The analysis was conducted under the following conditions: sample injection volume, 10 pL; UV detection, 210 nm; flow rate, 0.6 mL / min; column temperature, 40°C.

[0063] Confocal microscopy

[0064] Intracellular polyhydroxybutyrate (PHB) granules in Cupriavidus necator were visualized using confocal laser scanning microscopy.54Bacterial cultures were collected by centrifugation, washed twice with sterile phosphate-buffered saline (PBS, pH 7.0), and resuspended in PBS to an OD600of 30. For fluorescence staining, 3 pL bacterial suspension was mixed with 0.3 pL Nile Red solution (1 mg / mL in dimethylsulfoxide) and incubated for 1 hour at room temperature. Twenty pL of a 2% (w / v) low melting agarose solution (preheated to 60°C) was applied for fixation, followed by 1 pL of the stained cell suspension. Fluorescence imaging was performed using a Zeiss LSM880 confocal laser scanning microscope (Zeiss, Baden-Wtirttemberg, Germany). PHB granules stained with Nile Red were visualized using an excitation wavelength of 514 nm, with fluorescence emission recorded in the range of 540-750 nm.

[0065] PHB extraction13 B29-099-2WO

[0066] PHB extraction was performed following the protocol described by Wongmoon and Napathorn with slight modifications.51PHB extraction was performed using a shaking water bath method with 1,3-dioxolane as the extraction solvent. A 5% (w / v) wet cell suspension was prepared in 2 mL of 1,3-dioxolane and transferred to a toed glass tube sealed with a butyl rubber stopper and aluminum crimp. The extraction was conducted at 80°C for 6 h in a shaking water bath at 100 rpm to ensure efficient polymer dissolution. Following extraction, PHB was recovered by adding three volumes of water to the solution, which induced phase separation. The PHB-enriched phase was collected by centrifugation at 4,000xg for 5 min at room temperature, and the harvested PHB was washed twice with water to remove residual solvent. The purified PHB was subsequently dried in an oven at 60°C for 3 days to obtain the final powder.

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[0122] Supplementary Note 1. Target current density

[0123] Here, we estimate the production rate of CO2-derived products required to supply a suitable amount of feedstock to a bioreactor. In our study, we sought to produce polyhydroxybutyrate (PHB) by converting CO2 into acetate using a membrane electrode assembly (MEA) system and subsequently upgrading acetate to the biopolymer with Cupriavidus necator.

[0124] We first calculate the theoretical carbon yield for acetate-to-PHB conversion according to the following chemical equation.19nCH3COOH 4(C4H6O2)n + 6nH2O + 2nCO2(eq. SI)

[0125] The input carbon is 9n 2 moles of carbon q Input carbon = — mol of acetate x - - - - -mole of acetate X 12 - mol carb -on (eq. S2)= 54n g of carbon

[0126] The output carbon is4 moles of carbon output carbon = n mol of PHB x x mole of PHB12— mol carb —on (eq. S3)= 48n g of carbon

[0127] We can calculate the theoretical yield of PHB carbon by dividing the output carbon by the input carbon:48 n g of carbon in PHBTheoretical carbon yield = - - - - - - = 88% (eq. S4)54 n g of carbon in acetate

[0128] The practical production of commodity chemicals generally targets 80% of the theoretical carbon yield.3In this study, we assume a 70% carbon yield in the bioreactor, which corresponds to a yield (g of PHB / g of acetate) of 50%. Furthermore, the microbial productivity required for commercialization is typically 2-4 g L1h1.2-419 B29-099-2WO

[0129] We estimate the required production rate of acetate to meet these targets using the following equation:Production rate of acetate (M-1)PHB productivity (g L~'1h-1) (eq. S5)Yield (g PHB / gaacetate) X 59.044 - mol, acetat.e

[0130] The necessary acetate production is 67-135 mM h"1, as calculated in eq. S5. In our system, the acetate stream is delivered as a mixture of acetate and anolyte. We adjust the anolyte volume to 5 mL per cm2active area, which yields a target current density (mA cm'2) in a range that maintains both an acceptable full-cell potential and high selectivity (~0.2 M acetate collection).

[0131] We use the following equation to calculate the current density for converting CO2 to acetate:Reaction rate (A cm"2)= Production rate of acetate (mol L~ h-1)8 moles of electron < ox, X Anolyte volume (L cm ) X — - - - - -'1 mole of acetate. hX 96485 C mol1X - 3600 s

[0132] with assuming the anolyte volume of 5 mL cm"2active area where the electrochemical reaction takes place.

[0133] The calculated reaction rate is 72-145 mA cm"2for the microbial production rate of 2-4 g L"1h"1with the 50% yield. In practice, lab-based bioreactors often operate at -30% yield, and the minimal anolyte volume may affect both full-cell potential and selectivity over extended operation.

[0134] Supplementary Note 2. Carbon footprint analysis

[0135] We assessed the carbon footprint of the two-step system for biopolymer production under different energy efficiencies of the CO2 electrolyzer and renewable electricity sources. The majority of energy consumption arises from CO2electrolysis, while the bioreactor has minimal heat and power demands - consuming only -3% of the electricity required for electrolysis.2However, CO2 is emitted at 0.25 g CCWg polymer during microbial fermentation (eq. SI), thus, we add this CO2emission to the total carbon footprint. If a CO2capture unit is included alongside the CO2electrolyzer and bioreactor, this emitted CO2can be captured.

[0136] The electrolysis energy cost is calculated using the following equation:20 B29-099-2WOElectrolysis energy (J g1acetate)8 moles of electron „ 1= —: -; - 7 - X 96485 C mol1X — - — - 1 mole of acetate59 g mol1acetate (eq. S7)1X^full~cel1 XSelectivity (%)

[0137] The carbon footprint is calculated using the following equation:Carbon footprint (g C02eg-1Polymer) = (Electrolysis energy +0.03xElectrolysis energy (j g~racetate) \., Z-1 - - I X Z.oE -7 kWh]Yield(f) / (eq. S8)x carbon intensity of renewable electricity (g C02ekWh-1)+ 0.25 g CO2eg- Polymer

[0138] We present three scenarios in Table SI: 1) 10% energy efficiency at 3.5 Vfau-ceib 400 mA cm’ and 30% Faradaic efficiency for C2 oxygenates, 2) 20% energy efficiency at 3.2 VM.Ceii, 400 mA cm’2and 56% Faradaic efficiency for C2oxygenates and 3) lab data with different renewable electricity sources including wind, hydro and solar electricity.5’6The yield is considered at 50% in Table SI. The results illustrate how the performance of CO2electrolyzer affects the overall carbon footprint of the produced biopolymer.1 2 Lab data Electrochemical performance metricsVfuJl-cel] 3.5 V 3.2 V 3.9 V Current density 400 mA cm’2400 mA cm’2400 mA cm’2Faradaic30% 56% 26% efficiencyEnergy10% 20% 7% efficiencyElectrolysis 153 GJ / tonne of 75 GJ / tonne 191 GJ / tonne of energy acetate of acetate acetate Carbon emission from the bioreactor (kg CO2e / kg polymer)0.25 0.25 0.25 Carbon footprint (kg CO2e / kg polymer)Wind electricity 1.21 0.72 1.48 Hydro electricity 2.31 1.26 2.89 Solar electricity 3.66 1.92 4.64

[0139] Table SI. The carbon footprint of the CO2 electrolysis systems with projected metrics and lab data.

[0140] Supplementary Note 3. Theoretical CO2consumption

[0141] Assuming complete conversion of CO2into either acetate or ethanol, the minimum CO2flux is calculated using the following equation.21 B29-099-2WOCO2flux (mol s-1cm-2)Current density (A cm-2) 1 mole of product96485 C mol-1n moles of electronsm moles of CO2X -: - - -: - 1 mole of productX _ 1Theoretical C02conversion efficiency (%)

[0142] Here, n is the number of electrons required for the CO2reduction reaction (CO2RR): 8 moles of electrons are required to convert 2 moles of CO2 into acetate, and 12 moles of electrons are required for CO2-to-ethanol conversion, m represents the number of CO2moles that are involved in CO2RR: 2 moles of CO2are required for both CO2-to-acetae and CO2-to-ethanol. A theoretical CO2conversion efficiency is 36% for CO2-to-acetate and 25% for CO2-to-ethanol in the alkaline CO2electrolysis system.7

[0143] The minimum CO2flux for 8e’ products and 12e’ products is 2.1 pmol s'1cm’2and 2.1 pmol s’ 1 cm-’2 at 300 mA cm’ 2, respectively. At 400 mA cm -’2, CO2flux is 2.9 pmol s -’ 1 cm -’2 and 2.8 pmol s’1cm’2. In experiments, < 1% of CO2is detected at the cathodic outlet at < 2.1 pmol s’ and applied current density of 300 mA cm’.

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[0150] 7. Huang, J. E.; et al, CO2electrolysis to multicarbon products in strong acid.Science 2021, 372 (6546), 1074-1078.22 B29-099-2WO

Claims

CLAIMS1. A method for upgrading CO2 into a biopolymer comprising using a two-step abiotic-biotic system, comprising an electrolysis system, wherein CO2 is converted into C2 oxygenates (such as acetate and ethanol) at the cathode, using a Cu-Ag tandem electrocatalyst, the electrocatalyst comprising a membrane electrode assembly (MEA) cell, the cell comprising metal plates, an anion exchange membrane (AEM), a cathode and anode, wherein the cathode comprises nanoscale Cu and Ag nanoparticles on a hydrophobic carbon substrate, wherein CO2 is fed to the cathodic side and converted to C2 oxygenates at the electrocatalyst surface, wherein the electrolysis process generates a liquid stream containing oxygenates in a bio-compatible electrolyte (anolyte), wherein these electrosynthesized oxygenates are then fed to a bioreactor, where substrates are upgraded to the biopolymer.

2. The method of claim 1, wherein the biopolymer is 3-polyhydroxybutyrate (PHB).

3. The method of claim 1, wherein the biopolymer is 3-polyhydroxybutyrate (PHB), wherein PHB is biosynthesized from acetate in a Cupriavidus necator (C. nectar) bioreactor, wherein acetate is taken up by C. necator and converted to acetyl-CoA via acetyl-CoA synthetase (acs), wherein acetyl-CoA enters the TCA cycle for energy generation or serves as a precursor for PHB biosynthesis, wherein in the PHB biosynthetic pathway, PhaA catalyzes the condensation of two molecules of acetyl-CoA to form acetoacetyl-CoA, which is then reduced to 3-hydroxybutyryl-CoA by PhaB. Finally, PhaC polymerizes 3-hydroxybutyryl-CoA into polyhydroxybutyrate (PHB).

4. The method of claim 1, wherein the C. nectar is engineered to express genes encoding enzymes involved in PHB biosynthesis.

5. The method of claim 1, wherein the system is configured substantially as disclosed in Fig.1A. 3C, 4A or 4B.

6. The method of claim 1, wherein the system is configured as disclosed in Fig. 1 A, 3C, 4A, and 4B.

7. The method of claim 1, wherein the system is configured to produce CC>2-to-C2 oxygenates for fuel synthesis, or bioproducts synthesis in combination with biological reactions.

8. 1'he method of claim 1, wherein the system is configured with formulations herein, or with CO2 to C2 efficacies wherein, especially for liquid C2 products.23 B29-099-2WO

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