Green Chemistry View Article Online Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. PERSPECTIVE Cite this: DOI: 10.1039/c7gc01801k View Journal An overview of cathode materials for microbial electrosynthesis of chemicals from carbon dioxide Nabin Aryal, a,b Fariza Ammam, c Sunil A. Patil *†d and Deepak Pant *e The applicability of microbial electrosynthesis (MES) for chemical synthesis from carbon dioxide (CO2) requires improved production and energetic eﬃciencies. Microbial catalysts, electrode materials, and reactor design are the key components which inﬂuence the functioning of such processes. In particular, cathode materials critically impact the electricity-driven CO2 reduction process by microorganisms. Interest in cathode surface modiﬁcations for improving MES processes is thus consistently increasing. In this paper, the recent developments and spatial modiﬁcation of cathode materials for microbial CO2 reduction are systematically reviewed. The characteristics of commercially available materials, their modiﬁcations, and developments in new materials that have been used as cathodes for MES are summarized. Key cathode–microorganism interactions that led to improved CO2 conversion are then discussed. The cathode surface modiﬁcation approaches have focused mainly on improving the surface area and surface chemistry of the materials. Although the modiﬁed cathode surfaces improved bioﬁlm growth in direct electron uptake based bioconversions, they have achieved lower acetate production rates than that Received 17th June 2017, Accepted 5th October 2017 DOI: 10.1039/c7gc01801k rsc.li/greenchem 1. of hydrogen-based MES processes thus far. Research eﬀorts on diﬀerent materials suggest that the threedimensional cathodes that can retain more biomass, in particular in hydrogen-based bioconversions, are promising for further improvements in production eﬃciencies. Further eﬀorts toward reducing the energy inputs for achieving energetically eﬃcient MES processes by using electrocatalytically eﬃcient cathodes are needed. Introduction Microbial electrosynthesis (MES) is an emerging novel green technology for the production of biochemicals and biofuels from CO2 using energy sources derived from a poised cathode1–3 (Fig. 1). The energy can be supplied from renewable sources such as solar, wind, and biogas to catalyze CO2 reduction at the cathode by autotrophic acetogenic bacteria.4 MES thus opens up an alternate sustainable route for eﬃcient storage of surplus electrical energy into chemical bonds of easily transportable gas or liquid fuels or chemical a Biological and Chemical Engineering, Aarhus University, Hangovej 2, DK-8200 Aarhus N, Denmark b Danish Gas Technology Centre, Dr Neergaards Vej 5B, DK-2970 Horsholm, Denmark c Department of Engineering Science, University of Oxford, Parks Road, Oxford, OX1 3PJ, UK d Institute of Environmental and Sustainable Chemistry, TU Braunschweig, Hagenring 30, 38106 Braunschweig, Germany. E-mail: email@example.com e Separation and Conversion Technology, Flemish Institute for Technological Research (VITO), Boeretang 200, Mol 2400, Belgium. E-mail: firstname.lastname@example.org, email@example.com † Present address: Department of Earth and Environmental Sciences, Indian Institute of Science Education and Research (IISER), Mohali, Sector 81, S A S Nagar, Punjab 140306, India. This journal is © The Royal Society of Chemistry 2017 commodities.5 When coupled to the photovoltaic system, this essentially becomes an artificial photosynthesis device mimicking the CO2-to-chemicals process, where the conversion eﬃciency is up to 10% higher than natural photosynthesis bioprocess based bioenergy systems.6,7 Electrons can also be supplied from the anodic substrate oxidation process in a so-called microbial fuel cell (MFC) through an external circuit.8 Autotrophic acetogenic microorganisms utilize the acetylCoA/Wood–Ljungdahl (WL) pathway to fix CO2.9 MES from CO2 relies on the electron uptake capability of these microorganisms from the cathode. Microbes can either utilize electrons directly from the cathode or use reduced mediators or regenerated equivalents from the electrode as the electron source.10 The molecular mechanisms involved in cathodic electron uptake processes remain still poorly understood.11 So far, limited microorganisms for cathodic CO2 conversion have been identified through either screening of pure homoacetogens (mostly Sporomusa and Clostridium spp.) or enrichmentselection from the mixed culture inoculum sources. The first report on CO2 conversion to organic compounds such as methane and acetate using microorganisms in bioelectrochemical reactors dates back to the mid-1990s.12 Later, in a pioneering study, the possibility of providing the acetogenic Green Chem. View Article Online Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. Perspective Fig. 1 A schematic of the MES for CO2 reduction to products on a biocathode. microorganisms such as Sporomusa and Clostridium spp. with electrons delivered directly from a cathode to synthesize acetate from CO2 was reported.1,4 Since then, the production of a wide range of other chemicals, such as formate, butyrate, ethanol, isopropanol, and butanol has been reported in CO2-fed bioelectrochemical reactors.13–17 Another potential approach in the power-to-gas and CO2 conversion framework is electromethanogenesis, which leads to the production of methane gas.18–20 This production pathway is not considered in this paper as it has been recently reviewed in detail.18,21 The major bottlenecks for the development of MES technology as an economically feasible CO2 utilization approach are poor production indicators such as low production rates, low Dr Nabin Aryal is a post-doctoral researcher at Danish Gas Technology Centre (DGC) and Aarhus University Denmark. He received his M.Sc. degree in Environmental Technology and Engineering from Ghent University, Belgium, with an Erasmus Mundus fellowship and then a Ph.D. degree from the Technical University of Denmark (DTU) in Environmental Biotechnology. Currently, his Nabin Aryal research focuses on the optimization of bioelectrochemical cells for methane production, biogas upgradation and carbon dioxide utilization from gasifiers. He has 6 peer-reviewed publications and >95 citations (h-Index 4). Green Chem. Green Chemistry product titers, and energetic eﬃciencies. Microbial catalysts, the target product, and electrode materials mainly determine these key performance indicators. Diﬀerent approaches have been deployed in the recent years to optimize the MES process and improve these production indicators. These include, e.g., the manipulation or augmentation of cultivation medium composition to provide suﬃcient nutrients, improving reactor design and feeding conditions, enrichment of mix culture inoculum, operating reactors at optimized conditions and the spatial arrangement of the cathode to enhance its interaction with microorganisms.22–26 The cathode, by acting as an electron donor source to microbes, critically determines the MES performance. Desired electrode materials for use as biocathodes should have the following properties: (i) high conductivity, (ii) excellent chemical stability, (iii) high mechanical strength, (iv) biocompatibility, (v) high surface area and (vi) low cost.27–29 Recently, an increasing number of research eﬀorts have been devoted toward cathode material development and spatial surface modification to achieve MES productivity enhancements and bring it closer to its counterpart fermentation technology. Acetate, being the most widely investigated and most prominent product of autotrophic CO2 fixation, is often considered for evaluating the cathode performances in MES systems.14 In this review, we provide up-to-date information on the cathode materials used for CO2 conversions to liquid chemicals (mainly acetate) via the MES route in bioelectrochemical systems (BESs). Firstly, the commercially available materials used as cathodes in MES processes are described. This is followed by a discussion on the spatial modifications of the materials and purposely built cathode materials. The key interactions between bacteria and cathodes that lead to enhanced bioproduction are then discussed. Finally, future research perspectives and directions for advancing MES as a sustainable CO2 utilization approach are provided. Fariza Ammam Dr Fariza Ammam is a postdoctoral researcher at the Engineering Department, University of Oxford. She received her M.S. degree in Microbiology and a Ph.D. degree in Molecular Microbiology from the University of Paris Sud XI. Her main interests are microbial physiology, bioenergetics and synthetic biology for anaerobes. She is currently working on engineering and optimising CO2fixing cell factories for the bioproduction of fuels and chemicals in bioelectrical systems. This journal is © The Royal Society of Chemistry 2017 View Article Online Green Chemistry Nevin et al. (2010) described the first proof-of-principle of direct CO2 reduction by using a negatively charged solid-state graphite block cathode as an electron donor source.1 Since then, several commercially available carbon electrodes of diﬀerent shapes and types such as rod or stick, block, cloth, plate, activated carbon, gas diﬀusion activated carbon, granules, fiber rod, felt and reticulated vitreous carbon (RVC) have been commonly used as cathodes in MES systems (Table 1). Characteristics such as chemical resistance to corrosion, biocompatibility, and low-cost fabrication along with their proven applicability as bioanodes in several other BESs drive their wider and regular usage as cathodes in MES processes.23,25,30–32 In particular, graphite has become a predominantly used commercial carbon material. It is mainly used in the form of block or plate, rod or stick, and granules. It possesses a plane sheet structure and oﬀers intrinsic advantages in aqueous solution, such as a wide electrochemical window, relative inertness, suﬃcient electrical conductivity, low residual current, ease of modification, recyclability, reusability and very high biocompatibility.33 However, the use of a single graphite block or rod has a limitation to achieve higher productivity because of its low porosity and low accessible surface area for the adherence of microorganisms. Marshall and coworkers, therefore, proposed to use granular graphite as a cathode and achieved a high volumetric acetate production rate from CO2 with an enriched electrosynthesizing mix microbial culture community. 15,34 Electron micrographs confirmed increased bacterial cell attachment and growth on such cathodes over time. Such granular graphite materials provide a high volumetric surface area and porosity to favor electrode– microorganism interactions and sustain electrogenic biofilm activity.15,34 Other two-dimensional (2D) thin morphology carbon materials such as carbon plate and carbon cloth are also widely used owing to their better electrical conductivity, chemical stability, light weight, flexibility, and higher porosity compared to the graphite electrode. An early representative study by Zhang et al. used carbon cloth in MES systems due to its simplicity, flexibility and ease for further surface modification.29 Such plane 2-D electrodes, however, possess several disadvantages such as availability of low reactive specific surface area, low electrocatalytic activity, high internal resistance, high activation overpotential, and rapid creation of a passivation layer on the electrode surface thereby limiting their use in MES systems. Carbon felt and carbon fiber rod electrodes are examples of three-dimensional (3D) materials that have been explored in MES systems (Table 1). The three-dimensionally arranged topology of these electrodes oﬀers a high volume reactive surface area for the growth of biomass and minimizes mass transfer limitations. Furthermore, electrodes with 3D structure are flexible for further spatial modification, and their catalytic reactive sites oﬀer better interactions for electron exchange through either direct or mediated processes.35 Carbon felt has been widely used due to its high porosity, high surface area, and Dr Sunil Patil (Ph.D., Microbiology, 2011) is an Assistant Professor at Indian Institute of Science Education and Research (IISER), Mohali. His research interests include microbial electrochemistry, CO2 conversion to biochemicals and biofuels through microbial electrosynthesis, resource recovery from wastewaters using microbial fuel and electrolysis cells, and electro-bioremediation Sunil A. Patil of micropollutants. He has been a recipient of the DAAD scholarship (2008–2010), the Marie Skłodowska-Curie fellowship (2013–2015) and the Humboldt fellowship for experienced researchers (2016–2017). He has published 28 peer-reviewed publications (>1350 citations; H-index 21) and 8 book chapters. Dr Deepak Pant is a senior scientist at Flemish Institute for Technological Research (VITO) currently working on bioenergy, specifically, the design and optimization of microbial fuel cells (MFC) for energy recovery from wastewaters and microbial electrosynthesis (MES) for the production of value-added chemicals through electrochemically driven bio-processes. He has a Ph.D. degree in environDeepak Pant mental biotechnology and has 82 peer-reviewed publications (>4700 citations; h-index 36) and 22 book chapters to his credit. His research experience lies in bioelectrochemistry, microbial fuel cells, electrosynthesis, biofuels, bioenergy and life cycle analysis (LCA). He serves as the Editor for the new Elsevier Journal “Bioresource Technology Reports”. 2. Cathode materials used for MES from CO2 Carbon based materials are the most widely used cathodes for the bioconversion of CO2 in the MES systems. Fig. 2 depicts the timeline of the use of diﬀerent commercially available and surface-modified cathode materials in CO2 fed MES systems along with performance improvements (in terms of cathode surface based production rates) over the years. Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. Perspective 2.1 Commercial cathodes This journal is © The Royal Society of Chemistry 2017 Green Chem. View Article Online Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. Perspective Green Chemistry Fig. 2 Timeline depicting the developments of the cathode materials for CO2 reduction to acetate in MES systems. The highest acetate production rates (in terms of g m−2projected cathode surface area d−1) achieved in each year are presented in parentheses. For detailed information on diﬀerent other aspects of these MES studies, refer to Tables 1 and 2. SS – stainless steel, Au – gold, Pd – palladium, Ni – nickel, CNT – carbon nanotube, Si–TiO2 – silicon-titanium oxide, 3D RVC – three dimensional reticulated vitreous carbon, rGO – reduced graphene oxide, TEPA – tetraethylene pentamine, MWCNT – multiwall carbon nanotube. flexibility for further surface modifications. In a recent study, Bajracharya et al. optimized the acetate production rate by adding a stainless steel current collector to the carbon felt cathode and by lowering the applied potential to −690 mV versus SHE for in situ hydrogen generation to mediate electron transfer.31 Its use for the bioconversion of CO2 has also been successfully demonstrated in MES systems operated in a galvanostatic mode.25,36 In galvanostatic control, by applying a constant current, H2 can be generated continuously at cathodes for achieving non-limiting H2-based bioproduction. Recently, the direct use of unmodified RVC foam as cathodes has also been reported.37 In this study, the combination of galvanostatic control and the high surface area of the RVC cathode along with a continuous flow mode operation of biocathodes resulted in the high rate production of acetate (196.8 g m−2 d−1) from CO2. Stainless steel (SS) materials have been mostly used either as current collectors or in combination with the carbon-based cathodes in MES systems.25,31 Its direct ‘as is’ use without any surface modification is limited mainly due to its poor biocompatibility. However, its potential applicability as cathodes, in particular for H2-based bioproduction, has been demonstrated.38,39 2.2 Purposely built or modified cathodes The use of surface modified and purposely built materials have been increasingly proposed for biocathode optimization for MES from CO2 (Table 2). Before surface modification, electrodes are subjected to pretreatment, which potentially exposes the reactive surfaces for further modification and immobilization. The electrodes undergo either chemical treatment or thermal treatment to remove impurities from their surfaces. Commercially available carbon cloth and carbon felt are the most commonly used base materials for the spatial modification of electrodes because of their outstanding characteristics of flexibility, high porosity, high surface area for bacterial adherence and ease of spatial modification.23,29 For example, Zhang et al. modified carbon cloth electrodes by Green Chem. immobilizing chitosan, cyanuric chloride, 3-aminopropyltriethoxysilane, polyaniline, melamine, ammonia, gold, palladium, carbon nanotube (CNT) cotton and CNT polyester for CO2 reduction with S. ovata.29 Among the modified cathodes, chitosan, an amino and hydroxyl-group rich polysaccharide, increased the acetate production rate by 7.6-fold compared to its counterpart, unmodified carbon cloth cathode. This was attributed to its better biocompatibility resulting in improved electrostatic interactions between negatively charged bacterial cells and the positively charged chitosan-modified electrode. The catalytic surface with suitable pore size properties also enhanced biofilm growth on the cathode. Furthermore, carbon cloth modification with metal nanoparticles, such as gold (Au), palladium (Pd), or nickel (Ni) resulted in an increase of acetate production rates by 6-, 4.7- and 4.5-fold, respectively, compared to untreated electrodes. The low charge transfer resistances and the high conductivity of the nanoparticles were suggested to be at the origin of the enhanced acetate production in MES reactors.29 Similarly, cotton and polyester fabric based cathodes treated with CNTs also showed 3.4- and 3.2-fold higher acetate production rates compared to the untreated carbon cloth cathode. CNTs are one of the most promising materials for designing electrodes of bioelectrochemical devices, in particular, of biosensors and MFCs due to their high conductivity and biocompatibility but are relatively expensive.50,51 In another study by the same group, nickel (Ni) nanowires coated on a graphite stick with microwave treatment enhanced its surface roughness by almost 50 times compared to untreated graphite52 The porous structure of Ni-nanowire graphite substantially enhanced cathode–microorganism interactions leading to a 2.3 fold increased acetate production rate over untreated electrode.52 Jourdin et al. developed a NanoWeb-RVC cathode by using a chemical vapor deposition (CVD) method, which resulted in substantial enhancement of acetate production rate by 33.3 fold compared to the unmodified carbon plate electrode.27 Modified NanoWeb-RVC oﬀers a high surface area This journal is © The Royal Society of Chemistry 2017 View Article Online Green Chemistry Table 1 Perspective Commercial cathode materials used in CO2 fed MES systems Acetate production Cathode materials Microbial culture or inoculum source Operation mode Applied potential (mV vs. SHE) Carbon rod Anaerobic digestion slurry Sporomusa ovata S. sphaeroides S. silvacetica Clostridium aceticum Clostridium ljungdahlii Moorella thermoacetica Brewery wastewater Batch −201 −4.1 0.0431 0.2 CH4 Ng 12 Batch Batch −400 −400 −400 −590 1.38 0.062 0.045 0.006 0.14 0.104 Ng 0.063 0.0028 0.002 0.0027 0.0065 0.0065 1.7 2-Oxobutyrate — 2-Oxobutyrate — 2-Oxobutyrate, formate — H2, CH4 86 ± 21 84 ± 26 48 ± 6 53 ± 4 88 ± 2 85 ± 7 67 1 Batch −0.207 −0.017 −0.006 −0.024 −0.031 −0.031 Ng Brewery wastewater Batch −590 Ng Ng 10.5 H2, CH4 69 Batch −853 −953 −400 −3.3 −9.2 −0.03 16.3 19.2 0.063 0.33 0.39 0.02 24 40 18 35.2 ± 4.4 41 −1.5 −3.1 Ng −3.23 −5 12.8 1.58 9.75 38 19 0.127 0.11 Ng 0.34 1.29 81 56.6 89.5 40 ± 4 58 ± 5 39 42 Batch Batch −690 −703 −903 −600 −1260 H2, CH4 H2, CH4 Ethanol, H2, butyrate, propionate, & butanol — H2 H2 H2, formate H2 Batch −1140 −5 20.4 13.5 — 60.8 36 −1.7 −12.3 9.68 2.7 2.065c Ng H2 CH4 89 ± 12 28.9 44 45 Batch Batch −895 −690 −600 −690 −10 −1 −34.53 −2.1 40 7.51 18.35c 51.1 0.63 0.63 4.13 0.89 H2, CH4 and ethanol Ethanol, H2 Ethanol — 45 ± 7 31 64 ± 35 29.13 46 85.3 ± 8.3 26 Batch −800 −20b 92.54c 2.8c 16 Batch Batch Batch −660 −690 −690 −795 8.2c 33.28 8.1 14.41 3.04 10.4 10.7 16.57 0.244 0.1 0.41 0.73 0.18 0.53 0.54 10 53 87.6 ± 6.5 91.8 ± 5.3 61.1 ± 12 84.9 ± 4 69.9 ± 0.9 90.8 ± 12 50 38 22 30 Batch −1.4 Ng −0.45 −0.78 −0.19 −0.57 −0.7 −10 Ethanol, butyrate & butanol H2 Ethanol — — — — — H2, butyrate, ethanol Continuous −800 ∼−23 108 2.56 H2 33.6 48 Continuous −1100 to −1300 Continuous −1000 −83.3 196.8 3.6 H2 35 37 −5 21 3.5 Butyrate & isopropanol 63 17 −0.37 4.9 0.1 — 4.9 49 Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. Graphite stick Graphite stick Graphite granular Graphite granular Carbon felt Domestic wastewater sludge Carbon fiber rod Bog sediment SS felt Carbon felt Graphite rod Carbon felt Batch Acetobacterium woodii Batch Domestic WWTP sludge Batch Brewery wastewater Enriched mix culture (from UASB eﬄuents) Carbon felt with Enriched mix culture SS (from UASB eﬄuents) Graphite stick S. ovata Graphite Anaerobic digester granular sludge Carbon felt with Wastewater sludge Clostridium ljungdhalii SS Graphite stick Enriched mix culture Graphite stick Methanol adopted S. ovata Carbon cloth Enriched mixed culture from syngas SS plate S. ovata Graphite block S. ovata Graphite block S. ovata 2662 S. ovata 2663 S. ovata 3300 S. acidovorance S. malonica Graphite felt Enriched anaerobic with graphite sludge stick Carbon felt with Enriched anaerobic SS sludge RVC foam Enriched acetogenic culture Carbon felt with Enriched mix culture SS Carbon Paper S. ovata 2662 Batch −740 Continuous −600 Batch Batch −690 Maximum Current titer densitya Ratea (A m−2) (g m−2 d−1) (g L−1) Other products CE in acetate (%) Ref. 4 15 34 43 25 47 a Calculated based on projected surface area. b Maximum reported current density on the 34th day. c Approximate calculation from given production graph, WWTP – wastewater treatment plant, ng – not given, SS: stainless steel; CE: Coulombic eﬃciency. with macroporous size that enhances mass transfer and biofilm formation.27 These researchers further developed CNT modified RVC electrodes (referred to as EPD-3D) using an electrodeposition method and reported a high rate synthesis of acetate from CO2 with an enriched, stable acetogenic microbial community.58 They further modified RVC with multi-walled CNTs (MWCNTs) using the same technique. MES systems with This journal is © The Royal Society of Chemistry 2017 these cathodes resulted in an acetate production rate of up to 1330 g m−2 d−1, the highest to date based on the projected cathode surface area. This exceptionally high rate of bioproduction was achieved by utilizing highly open macroporous RVC with a pore size of 0.6 mm that resulted in a good balance between high surface area availability for biofilm formation and eﬃcient mass transfer between the biocathode and the Green Chem. View Article Online Perspective Table 2 Purposely built and surface modiﬁed cathode materials used in CO2 fed MES systems Cathode materials Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. Green Chemistry Carbon cloth + Chitosanb Cyanuric chlorideb 3-Aminopropyltriethoxysilaneb Polyanilineb Melamineb Ammoniab Goldb Palladiumb Nickelb CNT-cottonb CNT-polysterb Graphite stick – Ni nano wire Nanoweb 3D RVC 3D RVC with CNT MWCNT-RVC Activated carbon VITO-CoRE™d Si–TiO2 nanowire photocathode Carbon cloth-reduced graphene oxide tetraethylene pentamine (rGO-TEPA-CC) 3D-graphene carbon felt composite Gas diﬀusion activated NORIT® carbon 3D iron oxide modified carbon felt Graphene paper Microbial culture source S. ovata S. ovata WWTP sludge Enriched mix culture from WWTP sludge Same as above Mix culture Acetate production Applied Current potential densitya Maximum Fold CE in (mV vs. SHE) (A m−2) Ratea (g m−2 d−1) titer (g L−1) increase f acetate (%) Ref. −400 −0.47 −0.45 −0.20 13.51 ± 3.30 12.09 ± 2.95 5.6 ± 1.18 0.59e 0.79e 0.31e 7.6 6.8 3.1 86 ± 12 81 ± 16 82 ± 11 5.32 ± 1.29 1.8 ± 0.47 1.65 ± 0.82 10.67 ± 2.59 8.32 ± 2.06 8.024 ± 1.94 6.618 ± 1.47 5.66 ± 1.416 3.38 195 ± 30 685 ± 30 0.18e Ng −400 −850 −850 −0.18 −0.06 −0.06 −0.38 −0.32 −0.30 −0.22 −0.21 −0.63 −37 −102 0.35e 0.45e 0.23e 0.3e 0.13e 0.094 1.2 11 3 Nil Nil 6 4.7 4.53 3.4 3.2 2.3 33.3c — 85 ± 7 80 ± 15 82 ± 8 83 ± 14 79 ± 16 80 ± 15 83 ± 10 82 ± 8 82 ± 14 70 ± 11 100 ± 4 52 27 53 −1100 −400 −200 −0.165 1330 9.49 11 4.1 — — 84 ± 2 29.91 24 54 Ng 6 — 86 ± 9 55 29 S. ovata −595 −3.5 Methanol adapted S. ovata −690 −0.23 62.4 ± 26.64 1.88 11.8 83 ± 3 28 S. ovata −690 −2.4 54.57 ± 1.7 1.4 6.8 86 ± 3 23 Enriched Anaerobic sludge −1000 −20 36.6 2.89 — 35.46–88 56 e S. ovata −690 Ng 25.40e 1.8e 4.8 86 ± 9 57 S. ovata −690 −2.5 39.8 0.77 8g 90.7 49 a Calculated based on projected surface area. b Modified on the carbon cloth surface of electrode. c Compared to control carbon plate electrode. Trade mark plastic inert support electrode. e Approximate calculation from given production graph. f The fold increase is compared to the respective unmodified cathodes. CE – Coulombic eﬃciency, Ni – nickel, CNT – carbon nanotubes, Si–TiO2 – silicon–titanium oxide, 3D – Three dimensional, RVC – reticulated vitreous carbon, ng – not given, nil – negative result or no improvement. g Compared to control carbon paper electrode. d bulk electrolyte.59,60 In an interesting approach, Liu and coworkers developed the silicon-titanium oxide nanoparticle (Si-TiO2) photocathode to harvest sunlight directly. Using this photocathode, which harvested light as the sole electron source, up to 6 g L−1 acetate was produced with S. ovata.55 Recently, Cui et al. used iron oxide modified 3D carbon felt cathodes and reported 4.8-fold enhancement in the acetate production rate compared to the unmodified carbon felt cathode.57 The semi-conductive properties of ferric oxide were suggested to enhance the eﬃciency of extracellular electron transfer in this case. Graphene, whose use for the development and surface modification of diﬀerent non-conventional carbonaceous materials is on the rise,61 has also been used for developing cathodes for MES systems. For instance, Chen et al. functionalized tetraethylene pentamine in situ after reducing graphene on a carbon cloth electrode.28 In this case, the electric charge interaction accelerated biofilm formation and led to 11.8-fold enhancement in the acetate production rate compared to the Green Chem. unmodified electrode.28 Aryal et al. developed a 3D graphene carbon felt electrode for the formation of robust biofilm and reported enhanced acetate productivity by 6.8-fold compared to unmodified carbon felt.23 This research group further developed the freestanding and flexible graphene paper cathodes and reported 8-fold enhancement in acetate production compared to commercially available carbon paper electrode. The application of graphene electrodes49 in MES systems would be attractive considering its relatively low production costs.62,63 The use of other composite electrodes such as activated carbon VITO-CoRE® electrode and gas diﬀusion electrodes (GDEs) has also been investiagted.54,56 The porous composite activated carbon gas diﬀusion electrode provides an ideal three-phase interface (gas–liquid–solid). It consists of a hydrophobic gas diﬀusion layer to diﬀuse the CO2, a current collector, and a catalyst layer for the synthesis of chemicals by bacteria. The use of such a GDE led to an improved and controlled transfer of CO2 to the biocatalyst in MES as compared to the conventional gas sparged systems. The mass transfer coeﬃ- This journal is © The Royal Society of Chemistry 2017 View Article Online Green Chemistry Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. cient (kLa) for GDE was twice that of the sparged system.56 However, CO2 solubility was often limited by a thin diﬀusion layer formed with the wetted electrode surface.64–66 The same research group developed the VITO-CoRE™ electrode with plastic inert support as the cathode. This cathode with a high surface area led to enhanced biofilm formation and better productivity of MES from CO2 in single chamber reactors.54 3. Key cathode–microorganism interactions that lead to improved MES performance Electrode surface modification is an important strategy for enhancing electrode–microorganism interactions. Several reviews have summarized the required surface properties for engineering electrodes for bioanode applications.67–69 Some of the approaches reported for anode surface modification have also been employed successfully to generate better surfaces for cathodic processes. Biocompatibility and a high active surface area together with chemical stability are the main properties that have been taken into consideration for developing cathodes for MES processes so far.27–29,69 In microbial electrochemistry, projected surface area, Brunauer–Emmett–Teller (BET) surface area, biofilm covered area, electrochemically active surface and bioelectrochemically active surface have been used for the performance analysis of electrode materials.69 The BET surface is often used for reporting the performance of the modified electrode materials in MES systems. The consideration of bioelectrochemically active surface area in MES depends on the physical and biological interactions at the electrode materials. Perspective The key approaches used for the surface modification of cathodes are depicted in Fig. 3. These have mainly led to improving either the surface chemistry or the surface area or both, of the modified electrodes. A considerable improvement in the cathode surface based acetate production rates has been achieved by using surface modified or purposely built cathodes in MES systems. The key reasons or eﬀects that led to improvements in MES performance indicators are briefly discussed below. 3.1 Improved surface area The 3D electrodes having multiple layers of micro, meso and macro pores allow eﬃcient mass transfer and provide more surface to increase the bacteria loading (Fig. 3A). In this regard, a highly conductive 3D NanoWeb-RVC, developed by coating MWCNTs on RVC, generated a cathode with high surface to volume ratio optimal for bacterial attachment and mass transfer within the biofilm.28 Another electrode developed by using electrophoretic deposition by coating CNTs on RVC followed by chemical treatment resulted in a highly conductive cathode that delivered 100% electron recovery compared to 70% obtained with the NanoWeb-RVC.27,58 All these modifications benefitted from a macroporous structure of RVC that possessed high porosity and surface area. Besides improved surface area, these modifications also imparted additional properties to the cathode. For instance, the deposition of CNTs was suggested to improve the electrochemical communication between the microbes and the cathode.58 Furthermore, the functionalization of MWCNTs improves the electrochemical communication between the microbes by chemical treatment and was suggested to improve the conductivity of the material. It has been reported that the higher BET Fig. 3 Approaches adopted for modifying cathode surfaces and resulting eﬀects on bioﬁlm growth enhancement due to (A) improved 3D surface area and porous structure, (B) improved 3D surface area and functional group attachment, and (C) positively charged surface. This journal is © The Royal Society of Chemistry 2017 Green Chem. View Article Online Perspective Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. active surface area on an electrode helps to reduce the ohmic loss by improving the current density.70 In one MES study, the BET surface area of the reduced graphene oxide (rGO) functionalized carbon felt was increased by 2.2 fold after the modification of the base electrode (Fig. 3B).23 In this case, this modification led to manifold improvement in the MES productivity. Increase in the specific surface area using Fe2O3 nanoparticle deposition on the carbon felt cathodes that resulted in improved MES performance has also been reported. 3.2 Improved cell attachment and biofilm formation Improved surface chemistry can have a positive impact on the adherence of bacteria by providing a better electroactive nanosurface through hydrogen bonding, electrostatic attraction, and van der Waals interaction.24,29,71 The bacterial community used as biocatalysts greatly impacts biofilm formation and the electron transfer process. Therefore, a comparison between pure culture and mix culture driven MES regarding biofilm formation would not be appropriate. Mix cultures have been reported to be more tolerant to environmental stress conditions and fluctuations, and have shown higher production rates (Tables 1 and 2), which would suggest that they show better bioelectrocatalysis and form a better biofilm on the cathode than pure cultures.16,31,58 Two independent studies with 3D electrodes by Jourdin et al. highlight the role of modified materials in enhancing biofilm formation by the mix culture.27,58 In these studies, the same inoculum was used to allow the comparison between the eﬃciencies of diﬀerent materials including graphite plates, carbon felt, NanoWebRVC and EDP-3D. A well-developed and uniform biofilm with multilayers of microorganisms (thickness in the range of 5–10 µm) associated with a manifold increase in MES productivity was reported for the EDP-3D electrode. However, no uniform biofilm coverage was observed in the case of other electrodes inoculated with the same inoculum. MES with pure cultures in diﬀerent studies can be compared if the same microorganism and the same cathode potential were used. The pure cultures of many acetogens tested in MES systems including S. ovata, which represents the most well-studied electroautotroph so far, showed poor biofilm growth on unmodified graphite, carbon felt or stainless steel cathodes.1,4 Several initial reports on MES from CO2 described non-uniform, dispersed, patchy monolayers of cells on the poised cathodes.1,4,31 Among the numerous surface treatments applied by Zhang et al. for engineering a positively charged cathode only carbon cloth coated with cyanuric chloride or chitosan increased acetate production by 6 to 7 fold compared to the untreated electrodes.29 The confocal laser scanning microscopy images of the biocathodes showed nine times higher cell density biofilm on the chitosan modified cathode, thereby most likely supporting the higher acetate rate observed with this modified cathode.29 The authors argued that the Gram-negative bacterium S. ovata surface is negatively charged and the untreated carbon felt is neutral. Thus, the treatments applied had the objective to promote bacterial attachment and biofilm formation by means of adding positively charged Green Chem. Green Chemistry molecules on the electrode surface (Fig. 3C). Similarly, Chen et al. used rGO and positively charged tetraethylene pentamine nanoparticles to modify carbon cloth for improving cell attachment and reported a highly structured biofilm with the presence of what they named as self-assembled spheres formed by an adapted strain of S. ovata to methanol.28 3.3 Improved (bio)electrocatalysis Although the cathodic bioconversion processes have gained an increasing interest during the recent years, very little is known about the mechanistic of electron uptake by bacteria from a poised cathode.10,72,73 Extracellular electron transfer (EET) mechanisms from the cathode to the microbes have been hypothesized based on the electron transfer from the microbes to the anode.74 The cathodic EET may occur via two diﬀerent mechanisms depending on the microorganism and other experimental conditions involved: (i) mediator-free transfer (direct EET) or (ii) mediator-dependent transfer (indirect EET) (Fig. 4). In direct EET (Fig. 4A), a physical connection between the extracellular structures of the microorganism and the cathode is required. A few acetogenic bacteria like C. ljungdahlii which do not have cytochromes have been reported to be able to receive electrons directly from a poised electrode.4 The ability of the most commonly used homoacetogen, S. ovata, to achieve DET is suggested to not necessarily depend on the presence of specialized structures like nanowires but rather on the quantity of cellular redox active enzymes released under specific operating conditions. This is likely a realistic explanation for electron uptake by S. ovata in the absence of any redox mediators or energy carriers in MES processes. In fact, most of the operating conditions applied so far in pure culture driven MES systems, consisted of an additional step of pre-establishing biofilm on the cathode. This step requires growing bacteria with hydrogen for several days and may result in cell lysis and enzyme release that can eventually mediate the electron transfer. No direct contact with the solid surface is required for indirect EET. The indirect EET requires diﬀusible extracellular compounds such as redox mediators and energy carriers like H2 and formate that transport electrons between the electrode and the microorganisms. Enzymes like hydrogenases, and formate hydrogenases secreted by the biofilm matrix and released due to the cell lysis, have been identified to mediate indirect EET, for instance, in the case of identical processes with the electromethanogenic archaeon Methanococcus maripaludis.63,75 Like in direct EET, the mechanisms of mediatordependent cathodic EET are also still unclear (Fig. 3B and C). In the case of H2-mediated MES processes, a considerable lowering of the H2 evolution overpotential by biocathodes has generally been observed.24,25,37 This eﬀect has been attributed mostly to the biocatalytic activity of the microbial community grown on the cathodes. The role of modified cathode surfaces on enhancing the H2 electrocatalysis has not been reported and thus remains unclear in MES studies. Most of the studies on MES from CO2 with modified materials reported enhanced cathode–microorganism inter- This journal is © The Royal Society of Chemistry 2017 View Article Online Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. Green Chemistry Perspective Fig. 4 Proposed extracellular electron transfer mechanisms between the cathode and the microorganisms. Direct electron transfer may occur via cytochromes and/or nanowires (A), and indirect electron transfer may occur either by hydrogen mediation or enzyme mediation (B–C). actions but did not shed more light on the electron transfer mechanisms. However, these studies highlighted the impact of better engineered surfaces on biofilm development and enhanced bioelectrocatalysis. In this light, it can be speculated that independent of the electron transfer mechanism, which may be direct and/or indirect, biofilm growth and probably its thickness and heterogeneity play major roles in electron transfer eﬃciency. A control experiment without microbes has been performed to explain whether product formation has the direct contribution from the abiotic electrocatalytic activities of the electrode.49 None of the control experiments in MES has identified the electrochemical reduction of CO2 in aqueous solution. Hence, it can be concluded that the bioelectrocatalytic activity was principally responsible for the CO2 reduction process in MES systems. It is important to note that MES studies employ mostly carbon-based and SS materials as the cathodes. The electrochemical reduction of CO2 into hydrocarbons and other products has been reported on copper electrodes.76 In a pioneering study, Hori et al. reported the electrochemical formation of CH4, C2H4, EtOH and PrnOH by using single crystal electrodes and then described the molecular mechanism involved in this conversion.77 The surface morphology of the electrode had a critical role in the selectivity of product formation. Firstly, CO2 was reduced to CO2− by accepting an electron from the electrode and then it further reduced to CO. It was further proposed that CO was reduced to HCOO− at potentials more positive than −1.10 V vs. SHE.76–80 The mechanisms of CO2 reduction by biological catalysts diﬀer considerably. For example, several enzymes are involved in the conversion of two CO2 molecules into acetate via an acetyl-CoA pathway.9 As mentioned before, the electron uptake in biocathodes is either H2 mediated or direct microbe–cathode interaction based, depending upon the experimental conditions, in particular, the operational cathode potential and type of microbial catalysts used. The molecular mechanisms involved in electron This journal is © The Royal Society of Chemistry 2017 uptake processes at biocathodes are still not clear. The impact of the electrode surface properties on the biofilm structure and electron transfer mechanism is better understood and characterized in the case of bioanodes. For instance, Artyushkova et al. investigated the relationship between surface chemistry and the electrogenic biofilm by a multi-technique study combining analytical, spectroscopy, microscopy and electrochemical techniques. The aim of the study was to determine how functional groups like –N(CH3)3+, –COOH, –OH and –CH3 at the anode surface aﬀect the morphology, the chemistry and the functional properties of Shewanella oneidensis MR-1 biofilm.81,82 Positively charged and hydrophilic surfaces were found to be optimal for the growth of a uniform biofilm with very small cluster size and intercluster diﬀusion distance, which was suggested to be at the origin of the high electron transfer eﬃciency observed. The authors also stipulated that the biofilm heterogeneity is more important than the biofilm thickness. In contrary to what was expected, the influence of the substratum chemistry was found not only to aﬀect the initial cell attachment phase to the electrode surface but also to influence the cell–cell interactions and the biofilm architecture. In a diﬀerent study, Guo et al., investigated the eﬀects of surface charge and surface hydrophobicity on the anodic biofilm formation by using glassy carbon surfaces modified with –OH, –CH3, –SO3−, or –N + (CH3)3 functional groups. The results demonstrated that current output was correlated with the biomass quantity of the Geobacter population. Positively charged functional group attached surfaces and hydrophilic surfaces were more selective towards the electroactive microbes and thus formed more conducive electroactive biofilm.71 Recently, a model of mechanistic stratification occurring in the electroactive biofilm was described for G. sulfurreducens.83 According to this model, the nanowires coordinate with cytochromes present in the biofilm matrix to transfer electrons from the cells located in the upper layers of the biofilm to the cells located close to the anode. In this case, Green Chem. View Article Online Perspective Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. the nanowires directly discharge electrons from cells to cytochromes that act as gates between the biofilm and the anode. Similar mechanisms might also be speculated for the highly functionalized biocathodes obtained with diﬀerent surface modifications described in this review where the surface substratum improves the cell–cell interactions and the biofilm architecture, which in turn improves the electron transfer from the cathode to microbes. 4. Future perspectives The main challenge for MES processes is the performance improvement while maintaining low production costs. Multiple strategies such as enrichment of microbial inoculum, screening of new electroautotrophic microbes, reactor design, improving feeding conditions and cathode material modification have led to considerable improvements in CO2 conversion in MES systems. Key research considerations concerning cathodes and other components of MES systems that can be useful to advance this CO2 utilization approach further are presented below. • One of the limiting factors for the development of cathode materials is a lack of experimental understanding of electron exchange mechanisms from the cathode to the microbes. Moreover, a good understanding of the biofilm formation process on poised solid surfaces is necessary for further improvement of MES processes. In addition to the biocompatibility and high surface area, other parameters may favor the attachment of bacterial cells to the electrode surface. For instance, Philips et al. recently reported that C. ljungdahlii biofilm development on graphite and glassy carbon, two materials often used in MES, may be induced by NaCl addition.84 RNA-sequence analysis revealed the mechanistic of biofilm formation under stress conditions. Pilus-like appendages were observed and seen between the cells.84 Similar structures referred to as nanowires have been suggested to conduct electrons in the case of two electroactive bacteria Shewanella and Geobacter species.85,86 These findings on inducing biofilm formation on cathodes may provide a valuable approach for developing pure-culture driven MES processes. • Biofilm engineering for bioelectrocatalysis is as important as understanding the electron transfer mechanisms. The conductivity of the biofilm depends mainly on the physiological state of the cells within the biofilm. Due to the presence of diﬀerent strata of cells and depending on their location from the electrode, some cells in the biofilm may face respiratory limitation due to the lack of oxidised electron carriers. Indeed, a redox gradient has been reported to occur within the biofilm in an anodic process.83,87 Engineering biofilms with the right and precise thickness is crucial for the control of pH and nutrient gradients inside the biofilm matrix to ensure a uniform distribution of cell activity inside the biofilm. This will also contribute to maintaining the physiological state of the cells located in diﬀerent strata but also the redox and electrical gradients inside the biofilm.83,87 • The cost of cathode material development is one of the major bottlenecks toward approaching the practical appli- Green Chem. Green Chemistry cations for this process. Acetate production rates have been substantially improved in lab scale reactors; however, the pilot scale production has not been demonstrated yet. MES reactors with MWCNTs deposited on RVC and graphenebased cathodes have shown better performances, however, they are expensive. Recent analysis indicated that acetate production from MES is much more expensive than anaerobic fermentation technology due to the extra costs associated with the use of membrane and electrodes.88 Therefore, to make MES profitable, the recommendation was made on the development of high performance and durable electrode materials and membrane at minimum costs. Here, the knowledge gained in electrode development for other microbial electrochemical technologies can be explored for further development of cathodes for MES processes. • Achieving high current densities at the cathodes to realize a high rate production is a good strategy for enhancing MES productivity.23,58 Thus, the current density of the biocathode needs to be optimized by developing cheap and eﬃcient electrodes, for instance, for H2 production. Decreasing the H2 evolution overpotential of the cathodes by exploring metal-based, graphene, or composite materials is one option. So far, a limited number of metal-based composite electrodes has been applied in MES. These include mainly metals such as SS, iron, Pt, Au, and Ni.29,37 The experience of microbial electrolysis cells for H2 production might be applied to explore cathode materials for MES processes. The supply of H2 produced by electrolyzes to feed acetogens in fermenters might be an alternative strategy that needs to be explored further to achieve higher production rates.38 • More work on DET based bioproduction is necessary because the productivity is relatively low compared to the H2 mediated bioproduction in MES systems (Tables 1 and 2). This approach circumvents the washing out of the biomass from the systems, for instance in continuous reactors. As the bioconversions are facilitated at higher electric potential, this might also allow achieving higher energetic eﬃciencies thereby reducing the energy input for MES processes. • Optimization of operational conditions like improving electron supply turnover by galvanostatic control and continuous flow supply of growth medium to overcome nutrient limitations could warrant the productivity enhancement.25 A recent study by LaBelle et al. with controlled operating conditions resulted in a high rate acetate production.37 This study was performed with an unmodified RVC electrode previously reported to be not able to promote the biofilm development. In this case, the constant flow of fresh medium coupled to galvanostatic control resulted in 52-fold higher volumetric production rates compared to the one obtained with NanoWebRVC (0.78 g L−1 h−1 versus 0.015 g L−1 h−1, respectively).43,58 This represents the highest volumetric acetate production rate achieved thus far via the MES process, that too merely optimizing two operational conditions and without electrode surface modification. Consideration of other process parameters may also further enhance the MES process. For instance, CO2 is usually supplied by continuous bubbling in reactors, and due This journal is © The Royal Society of Chemistry 2017 View Article Online Published on 26 October 2017. Downloaded by University of Windsor on 26/10/2017 16:22:02. Green Chemistry to its low solubility in aqueous medium and its low mass transfer, only a small fraction of the total CO2 supplied during the process is utilized by the microorganisms. An eﬃcient gas supply with improved mass transfer is required for any real application considerations of MES. A gas diﬀusion electrode applied in MES by Bajracharya et al. has proved that appropriate gas supply may potentially lead to improved MES eﬃciency.56 • The anodic reaction on most of the MES systems has been water oxidation thus far. Further research should thus also be focused on identifying useful anodic conversions to make the whole MES system productive. • Importantly, further carbon chain elongation or product selection other than acetate is desired for developing economically attractive MES processes. The application of synthetic biology tools, metabolic engineering and adaptive evolution of acetogens might warrant targeting higher carbon chain compounds in MES.26 By applying a certain set of operational conditions, it is also possible to divert the production pathways, in particular, for mixed culture based MES processes, as recently reported.17 5. Conclusions This review summarizes the cathode materials that have been used in MES for the synthesis of chemicals from CO2. Along with the fundamental requirement of biocompatibility, it is essential to develop high surface area, highly porous and good conductive cathodes. Most of the strategies adopted to improve the cathode–microorganism interactions were targeted mainly toward biofilm enhancement. Biocompatibility and a high surface area were demonstrated to be required for developing uniform biofilms on the electrode, especially for pure culture biocathodes. The recent progress made with 3D electrode configuration seems to be the most promising material for MES processes. Further development of cheap and eﬃcient electrode materials will have a significant impact on the scaling-up potential of the MES technology. Undeniably, understanding the molecular mechanisms of electron transfer between a surface with low redox potential and microbial cells is necessary for further development of MES processes. Conﬂicts of interest The authors have no conflicts of interest to declare. Acknowledgements Nabin Aryal was supported by a FutureGas project from Innovation Fund Denmark – Innovationsfonden. Sunil Patil acknowledges the Alexander von Humboldt Foundation for the financial support. This journal is © The Royal Society of Chemistry 2017 Perspective References 1 K. P. Nevin, T. L. Woodard, A. E. Franks, Z. M. Summers and D. R. Lovley, mBio, 2010, 1, e00103–e00110. 2 D. R. Lovley and K. P. Nevin, Curr. Opin. Biotechnol., 2013, 24, 385–390. 3 S. A. Patil, S. Gildemyn, D. Pant, K. Zengler, B. E. Logan and K. Rabaey, Biotechnol. Adv., 2015, 33, 736–744. 4 K. P. Nevin, S. A. Hensley, A. E. Franks, Z. M. Summers, J. Ou, T. L. Woodard, O. L. Snoeyenbos-West and D. R. Lovley, Appl. Environ. 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