Anodic Biocatalyst Research Articles

Harvesting Bioelectricity from Microbial Fuel Cells (MFCs) Powered by Electroactive Microbes

The application of microbial fuel cells is still facing some challenges due to its low power output and high internal resistance. It is desirable to obtain a stable and consistent power output from an MFC to support practical real-world applications. Five electroactive bacteria (isolate LGf1, LGf11, LGf15, LGf20, and LGf22) isolated from the sediment of Waduk Saguling were exploited as the potential anodic biocatalyst for MFC, and the performance of these MFCs were studied in terms of voltage generation (open and close circuit), power density and the losses (polarization technique), and efficiencies (coulombic and energy). MFC biocatalyst by isolate LGf11 performed the best electrochemical performances, including highest OCV (open circuit voltage) value (804 mV) and power output (0.043 W/m2), lowest ohmic resistance (475 Ω), and highest coulombic efficiency (75.79%) and energy efficiency (88.36%) among all anodic biocatalysts. Nevertheless, all the five isolates were potential to be exploited as active biocatalyst for MFC due to their high OCV values and the stability of voltage generations, both in open circuit and close circuit mode. The development of system configuration and the use of more suitable substrate for different electroactive microbes in order to harvest more power output was recommended for further study. Utilization of these potential microbes for other applications in MFC (such as wastewater treatment etc.) was also suggested for further research. Keywords: Bio-electrochemical system, Biofuel, Efficiency, Electro-microbiology, Power output

Performance of Hybrid Process Constructed Wetland-Microbial Fuel Cell for Melanoidin Degradation and Simultaneous Electricity Generation

Melanoidin is the main cause of the dark brown color of the palm oil mill effluent (POME) that form under the Maillard reaction. In this study, the constructed wetland integrated with microbial fuel cell (CW-MFC) has been developed for melanoidin removal from the POME and simultaneously electricity generation as a by-product. The macrophyte Dieffenbachia sp. has been used as a biocatalyst on the cathode electrode and the oxidoreductase-producing bacterium Bacillus lichenformis with laccase and manganese peroxidase activity has been used as an anodic biocatalyst. The maximal melanoidin removal, chemical oxygen demand (COD) removal, enzyme activity, and power output were monitored. The maximal laccase and manganese peroxidase activities of 1.60 ± 0.10 U/mL and 1.45 ± 0.05 U/mL were found during melanoidin degradation. In addition, the maximal melanoidin removal of 93.59 ± 0.10% and 95.12 ± 0.15% were achieved respectively. When the maximal power density of 0.18 ± 0.01 mW/m3 was generated. This study gained new knowledge about using the CW-MFC system as a biological treatment process of the melanoidin content in the POME and simultaneously generated electrical energy as a by-product. HIGHLIGHTS The coupling process of constructed wetland and the microbial fuel cell is promising for the melanoidin removal of the POME The maximal melanoidin removal of 93.59 % was gained The maximal power output of 0.18 mW/m3 was generated from this system GRAPHICAL ABSTRACT

L-amino acids affect the hydrogenase activity and growth of Ralstonia eutropha H16

Ralstonia eutropha H16 is a chemolithoautotrophic bacterium with O2-tolerant hydrogenase (Hyds) enzymes. Hyds are expressed in the presence of gas mixtures (H2, O2, CO2) or under energy limitation and stress conditions. O2-tolerant Hyds are promising candidates as anode biocatalysts in enzymatic fuel cells (EFCs). Supplementation of 0.5% (w/v) yeast extract to the fructose-nitrogen (FN) growth medium enhanced H2-oxidizing Hyd activity ~ sixfold. Our study aimed to identify key metabolites (l-amino acids (l-AAs) and vitamins) in yeast extract that are necessary for the increased synthesis and activity of Hyds. A decrease in pH and a reduction in ORP (from + 240 ± 5 mV to − 180 mV ± 10 mV values) after 24 h of growth in the presence of AAs were observed. Compared to the FN-medium control, supplementation of 7.0 μmol/ml of the l-AA mixture stimulated the growth of bacteria ~ 1.9 to 2.9 fold, after 72 h. The whole cells’ H2-oxidizing Hyd activity was not observed in control samples, whereas the addition of l-AAs, mainly glycine resulted in a maximum of ~ 22 ± 0.5 and 15 ± 0.3 U, g CDW−1 activity after 24 h and 72 h, respectively. Our results suggest a correlation between ORP, pH, and function of Hyds in R. eutropha H16 in the presence of key l-AAs. l-AAs used in small amounts can be proposed as signaling molecules or key components of Hyd maturation. These results are important for the optimization of O2-tolerant Hyds production as anode biocatalysts.

Constriction of H2 biofuel cell voltammeter based on Escherichia coli microbial cells.

Hydrogen (H2) is the lightest and most abundant chemical element in the universe, with potential to replace fossil fuels as a carbon-free source of energy. Sensitive and faster H2 gas sensors are/will be required in many industries and hydrogen-based infrastructures. Microbial hydrogenases (Hyds) reversibly catalyze the formation and oxidation of H2. Isolation and characterization of O2-tolerant [NiFe]-Hyds have stimulated new concepts in hydrogen biofuel cells (BFCs). Platinum-independent H2-driven bio-electrochemical devices are suitable for environmentally friendly energy generation and application in biosensors. Escherichia coli employs four [NiFe]-Hyds, which can be applied as anode biocatalysts in BFCs. In this study, the H2 BFC voltammeter working with graphite based strips (sensors) were constructed and the efficiency of applying the E. coli BW25113 whole cells or crude extracts as anode biocatalysts were evaluated. E. coli BW25113 was grown under fermentation of 0.2% glucose or 1% glycerol in a peptone medium, pH 7.5. Upon glucose fermentation, H2-oxidizing Hyd activity of E. coli whole cells was 1.03±0.01 U (mg CDW)−1 and 1.39±0.01 U (mg CDW)−1 was observed in cell extracts. During glycerol fermentation, Hyd activity reached up to 2.15±0.02 U (mg CDW)−1 and 3.58±0.02 U (mg CDW)−1, respectively in whole cells and cell extracts. 3 μL volumes of bacterial whole cells or cell extracts were immobilized on the sensors with the aid of 3 mM KCl and 0.8% bacteriological agar support. Whole immobilized cells(grown on glucose) produced an electrical potential of 0.650±0.05 V, and with the cell extracts, the electrical potential was slightly lower. Meanwhile, under glycerol fermentation, the electrical potential was 0.550±0.05 V in cell extracts and whole cells. Despite, the highest electrical potential was achieved with bacterial whole cells grown on glucose, H2-oxidizing Hyd activity was stimulated upon bacterial glycerol fermentation.

Wolffia globosa as a biocatalyst in plant-based biofuel cells

The rootless duckweed Wolffia globosa, not explored toward electrogenicity till now, is investigated as a putative biocatalyst in Plant-based Biofuel Cells (P-BFC) for the electrical current generation and its basic metabolic changes during the polarization are depicted. After a short adaptation period, the open-circuit voltage of P-BFC, utilizing W. globosa as an anodic biocatalyst, reaches values of 630 mV. At a connected external resistor of 1 kΩ in the electric circuit, stable current densities of 170±10 mA m-2 are achieved. The electrical outputs depend on the anodic potential, reaching negative values of ca. -200 mV (vs. SHE). W. globosa produces an electrochemically active compound, acting as an electron shuttle. The polarization intensifies the W. globosa metabolism, expressed in a double increased glucose and starch content along with 1.82 times higher specific amylase activity of 70.0±2.8 U g-1 wet biomass in the organelle-enriched fractions of the explored as biocatalysts plants compared to the control. The results reveal that Wolffia globosa can be utilized as a biocatalyst in P-BFC for simultaneous electricity generation and increased carbohydrate and protein content.

A Biobattery Capsule for Ingestible Electronics in the Small Intestine: Biopower Production from Intestinal Fluids Activated Germination of Exoelectrogenic Bacterial Endospores

AbstractFunctioning ingestible capsules offer tremendous promise for a plethora of diagnostic and therapeutic applications. However, the absence of realistic and practical power solutions has greatly hindered the development of ingestible electronics. Microbial fuel cells (MFCs) hold great potential as power sources for such devices as the small intestinal environment maintains a steady internal temperature and a neutral pH. Those conditions and the constant supply of nutrient‐rich organics are a perfect environment to generate long‐lasting power. Although previous small‐scale MFCs have demonstrated many promising applications, little is known about the potential for generating power in the human gut environment. Here, this work reports the design and operation of a microbial biobattery capsule for ingestible applications. Dormant Bacillus subtilis endospores are a storable anodic biocatalyst that will provide on‐demand power when revived by nutrient‐rich intestinal fluids. A conductive, porous, poly(3,4‐ethylenedioxythiophene) polystyrene sulfonate hydrogel anode enables superior electrical performance in what is the world's smallest MFC. Moreover, an oxygen‐rich cathode maintains its effective cathodic capability even in the oxygen‐deficit intestinal environment. As a proof‐of‐concept demonstration in stimulated intestinal fluid, the biobattery capsule produces a current density of 470 µA cm−2 and a power density of 98 µW cm−2, ensuring its practical efficacy as a novel and sole power source for ingestible applications in the small intestine.

Small-scale, storable paper biobatteries activated via human bodily fluids

Herein, a fully paper-based biobattery composed of four microbial fuel cell (MFC) units is evaluated, one that can be prepared in advance, stored, and quickly activated with virtually any available fluid. The biobattery uniquely utilizes Bacillus subtilis endospores as the storable anodic biocatalyst; the dormant, robust nature of B. subtilis endospores should allow for device preinoculation with spores followed by prolonged storage of the fully fabricated paper battery until needed. A germinant paper layer strategically fabricated above the spore-loaded anode layer contains all of the necessary chemical germinants and nutrient components required for the endospores to begin germination, exit dormancy, and return to fully metabolic vegetative bacterial cells that can generate electrical energy. This mechanism allows for the battery to be simply initiated via a wide range of available liquids. Bioelectricity generation of the battery is successfully demonstrated after introduction of a variety of artificial bodily fluids, including saliva, sweat, and urine, along with tap water. Since the biobattery has the capability of serially linking all 4 of its MFCs through simple dynamic folding, the device’s total power output can be greatly enhanced; a single biobattery is able to achieve 0.56 V and 2.4 μW, which is beyond the ratings required for their intended application in single-use, disposable sensors. Therefore, this concept of integrating 4 spore-based MFCs into a single biobattery device with a built-in germinant layer offers a potential solution for stable, long-term storable power sources, displaying feasibility for integration with low power, disposable sensor applications.

Electrogenic degradation of Reactive Red 152 dye by Niallia circulans DC10 and its genome sequence analysis reveals genes mediating dye degradation and anodic electron transfer

This study demonstrates the application of a newly isolated bacterium N. circulans DC10 for the degradation of azo dye Reactive Red 152 (RR152) and bio-electricity generation in a microbial fuel cell (MFC) system. When used as an anodic biocatalyst, N. circulans DC10 achieved 94.78 ± 1.17% decolourization and 89.55 ± 2.28% COD reduction within 72 h in the MFC system along with 166.48 mW/m2 maximum power density generation. The FT-IR and GC–MS analysis suggested that N. circulans DC10 utilizes the phenylacetic acid catabolic pathway for degradation of RR152 in the dual chamber MFC system. The genome sequence analysis of N. circulans DC10 revealed the presence of flavin mononucleotide (FMN) dependant-NADH azoreductase gene azoR and a cluster of genes, most of which participate in the biosynthesis of type IV pili (T4P), an extracellular appendage. The RT-qPCR analysis showed that the expression levels of azoR gene increased around seven folds and T4P assembly genes pilT/pilZ expression levels increased more than two folds in N. circulans DC10 during active biodegradation of RR152 in the anode of MFC systems as compared to the static control flask cultures. The obtained results suggest that T4P has a role in promoting cell adherence to the surface of anode by N. circulans DC10 for exo-electron transfer generating bioelectricity. The cumulative effect of azoR and type IV pilus assembly genes resulted in efficient bio-electrogenic degradation of azo dye RR152 in the MFC system.

Gram-positive bacteria covered bioanode in a membrane-electrode assembly for use in bioelectrochemical systems

A novel strain of Gram-positive bacteria Paenibacillus profundus YoMME was recognized by sequencing of 16S rRNA gene and after that tested for exoelectrogenicity for the first time. It was found that at an applied potential of −0.195 V (vs. SHE) the bacteria are capable of generating electricity and forming electroactive biofilms for 3–4 days. A tendency for the decrease in double-layer capacitance and the increase in the charge transfer resistance during the maturation of the biofilm was established. The formed bioanodes were used as a part of a membrane-electrode assembly (MEA) together with a selected cathode (E-Tek) and a separator (Zirfon). The applicability of MEA with the bioanode was tested by operating a newly designed bioelectrochemical system in a microbial fuel cell (MFC) or microbial electrolysis cell (MEC) mode. A current density of 200 mA m−2 was generated by the MFC after the improvement of the cathodic reaction through facilitated air access. The Coulombic efficiency in different MFC runs ranged from 5.2 to 7.4%. It was also determined that 0.65 V applied cell voltage is appropriate for the operation of the cell in the electrolysis mode, during which a current density of 2–3 Am−2 was reached. This, along with the evolved gas on the cathode, shows that as an anodic biocatalyst P. profundus YoMME assists the electrolysis processes at a significantly lower voltage than the theoretical one (1.23 V) for water decomposition. The hydrogen production rate varied between 0.5 and 0.7 m3/m3d and the cathodic hydrogen recovery ranged from 49.5 to 61.5 %. The estimated energy efficiency based on the electricity input exceeds 100 %, which indicates that additional energy is being gained from the biotic oxidation of the available organics.

Recent trends in upgrading the performance of yeast as electrode biocatalyst in microbial fuel cells

Microbial fuel cell (MFC) is an optimistic fuel cell technology that applies microorganism's biochemical catalytic activities in consuming organic substrate and produce electricity. In the past, several researchers have reported power generation from Saccharomyces cerevisiae, but nowadays, most of the studies are centred around bacterial biofilms (prokaryotes) as anode biocatalyst. Yeast (a eukaryote) has also been applied as a biocatalyst in MFCs as they are non-pathogenic, easy to handle and tolerant to various environmental conditions. Yeast strains such as Arxula adeninvorans, Candida melibiosica, Hansenula polymorpha, Hansenula anomala, Kluyveromyces marxianus and Saccharomyces cerevisiae have been utilized in MFCs. This review summarizes the application of yeast as an anode biocatalyst together with a discussion on the mechanism of electron transfer from yeast cells to the anode and highlights the techniques applied in improving the efficiency of yeast-based MFCs. The recent challenges and benefits of utilizing yeast in MFCs have been also encapsulated in this review.

Electrospun Nanofibers for Applications in Energy Harvesting and Generation

Nanofibers have long been exploited as a versatile, attractive material compared to their bulk counterparts due to their exceptionally high specific surface area and aspect ratio, highly porous structure, and wide range of materials that can be used for synthesis. Such unique properties helped exhibit their potential in a variety of applications in areas of energy, healthcare, and environment. Applications in energy are widely studied especially because of the global need for a source of energy and method of energy production that is alternative to the combustion of fossil fuels, as it presents many environmental drawbacks, which ultimately lead to adverse health effects on humans. To bypass the use of fossil fuels, renewable sources such as wind, hydro, geothermal, and solar energy have been introduced; devices that take advantage of alternative sources of energy such as nanogenerators, fuel cells, and thermoelectric generators have been developed.In this work, hybrid nanofibers were fabricated with precise control of the working parameters to fine tune their properties for various applications, including piezoelectrics and enzymatic fuel cell electrodes. Pure polyacrylonitrile (PAN) nanofibers were first electrospun based on a series of design of experiments to minimize both the diameter and beading. By correlating the solution, electrospinning, and environmental conditions to the resulting solution and nanofiber properties, it was found that not only physical properties such as electrical conductivity but also electrochemical properties of the nanofibers could be controlled. Physical and electrochemical properties of porous carbonaceous nanofibers derived from PAN, zinc acetate and multi-walled carbon nanotubes were studied with SEM, 4-point probe measurement, CV, LSV, and EIS. Based on the electroanalytical methods, the best nanofiber mats were chosen and used as electrodes to immobilize multienzyme cascade systems, a concept of which was demonstrated with a glucose oxidase-BglA bienzymatic system on a customized DNA scaffold as the anodic biocatalyst.

Performance evaluation of yeast-assisted microalgal microbial fuel cells on bioremediation of cafeteria wastewater for electricity generation and microalgae biomass production

In this work, microalgae–microbial fuel cell (MMFC) was used as an alternative method to treat cafeteria waste in Indonesia to produce electricity and microalgal biomass simultaneously. The organic contents from cafeteria waste were used as substrates by Saccharomyces cerevisiae. Spirulina platensis acted as an oxygen producer for oxygen reduction reaction on the cathode side. A salt bridge was used as a separator and ion exchanger. Yeast (Saccharomyces cerevisiae) was used as an anode biocatalyst because of its availability, resistance to extreme weather, and ease of culture and incubation. A voltage of 0.17 V and current density of 400 mA m−2 were obtained using the MMFC system. Meanwhile, the power density and maximum power density (MPD) reached 73 and 98 mW m−2, respectively. The MMFC systems can easily reduce the chemical oxygen demand to 60% and the total dissolved solid to 82.83%. Meanwhile, the microalgae growing on the cathode side reached 0.74 g L−1 by utilizing CO2 from the anode side. The MPD of the MMFC system was 2.38 folds higher than that of the MFC system (without microalgae) and 1.58 times higher than that of the MMFC system that used a commercial membrane. These advantages make the MMFC system for cafeteria waste highly suitable for use in countries with large populations or developing countries.

Application of organic waste glycerol to produce crude extracts of bacterial cells and microbial hydrogenase-the anode enzymes of bio-electrochemical systems.

Glycerol is an organic waste material that can be used for the production of microbial biomass, consequently providing valuable biocatalysts promoting the generation of electrical current in microbial fuel cells (MFCs). [NiFe]-Hydrogenases (Hyds) of Escherichia coli and Ralstonia eutropha may be applied as potential anode biocatalysts in MFCs. In this study, E. coli K12 whole cells or crude extracts and R. eutropha HF649 synthesizing Strep-tagged membrane-bound Hyds (MBH) were evaluated as anode enzymes in a bioelectrochemical system. The samples were immobilized on the sensors with polyvinyl acetate support. Mediators like ferrocene and its derivatives (ferrocene-carboxy-aldehyde, ferrocene-carboxylic acid, methyl-ferrocene-methanol) were employed. The maximal level of bioelectrocatalytic activity of Hyds was demonstrated at 500mV voltage. Depending on the mediator and biocatalyst, current strength varied from 5 to 42 μA. Introduction of ferrocene-carboxylic acid enhanced current strength; moreover, the current flow was directly correlated with H2 concentration. The maximal value (up to 150 μA) of current strength was achieved with a 2-fold hydrogen supply. It may be inferred that Hyds are efficiently produced by E. coli and R. eutropha grown on glycerol, while ferrocene derivatives act as agents mediating the electrochemical activity of Hyds.

Characterization of pyranose oxidase variants for bioelectrocatalytic applications

Pyranose oxidase (POx) catalyzes the oxidation of d-glucose to 2-ketoglucose with concurrent reduction of oxygen to H2O2. POx from Trametes ochracea (ToPOx) is known to react with alternative electron acceptors including 1,4-benzoquinone (1,4-BQ), 2,6-dichlorophenol indophenol (DCPIP), and the ferrocenium ion. In this study, enzyme variants with improved electron acceptor turnover and reduced oxygen turnover were characterized as potential anode biocatalysts. Pre-steady-state kinetics of the oxidative half-reaction of ToPOx variants T166R, Q448H, L545C, and L547R with these alternative electron acceptors were evaluated using stopped-flow spectrophotometry. Higher kinetic constants were observed as compared to the wild-type ToPOx for some of the variants. Subsequently, the variants were immobilized on glassy carbon electrodes. Cyclic voltammetry measurements were performed to measure the electrochemical responses of these variants with glucose as substrate in the presence of 1,4-BQ, DCPIP, or ferrocene methanol as redox mediators. High catalytic efficiencies (Imaxapp/KMapp) compared to the wild-type POx proved the potential of these variants for future bioelectrocatalytic applications, in biosensors or biofuel cells. Among the variants, L545C showed the most desirable properties as determined kinetically and electrochemically.

Growth of the facultative chemolithoautotroph Ralstonia eutropha on organic waste materials: growth characteristics, redox regulation and hydrogenase activity

BackgroundThe chemolithoautotrophic β-proteobacterium Ralstonia eutropha H16 (Cupriavidus necator) is one of the most studied model organisms for growth on H2 and CO2. R. eutropha H16 is also a biologically significant bacterium capable of synthesizing O2-tolerant [NiFe]-hydrogenases (Hyds), which can be used as anode biocatalysts in enzyme fuel cells. For heterotrophic growth of R. eutropha, various sources of organic carbon and energy can be used.ResultsGrowth, bioenergetic properties, and oxidation–reduction potential (ORP) kinetics were investigated during cultivation of R. eutropha H16 on fructose and glycerol or lignocellulose-containing brewery spent grain hydrolysate (BSGH). BSGH was used as carbon and energy source by R. eutropha H16, and the activities of the membrane-bound hydrogenase (MBH) and cytoplasmic, soluble hydrogenase (SH) were measured in different growth phases. Growth of R. eutropha H16 on optimized BSGH medium yielded ~ 0.7 g cell dry weight L−1 with 3.50 ± 0.02 (SH) and 2.3 ± 0.03 (MBH) U (mg protein)−1 activities. Upon growth on fructose and glycerol, a pH drop from 7.0 to 6.7 and a concomitant decrease of ORP was observed. During growth on BSGH, in contrast, the pH and ORP stayed constant. The growth rate was slightly stimulated through addition of 1 mM K3[Fe(CN)6], whereas temporarily reduced growth was observed upon addition of 3 mM dithiothreitol. The overall and N,N′-dicyclohexylcarbodiimide-sensitive ATPase activities of membrane vesicles were ~ 4- and ~ 2.5-fold lower, respectively, upon growth on fructose and glycerol (FGN) compared with only fructose utilization (FN). Compared to FN, ORP was lower upon bacterial growth on FGN, GFN, and BSGH.ConclusionsOur results suggest that reductive conditions and low ATPase activity might be signals for energy depletion, which, in turn, leads to increased hydrogenase biosynthesis to overcome this unfavorable situation. Addition of fructose or microelements have no, or a negative, influence on hydrogenase activity. Organic wastes (glycerol, BSGH) are promising carbon and energy sources for the formation of biomass harboring significant amounts of the biotechnologically relevant hydrogenases MBH and SH. The results are valuable for using microbial cells as producers of hydrogenase enzymes as catalysts in enzymatic fuel cells.

High-Efficiency Enzymatic Fuel Cell Via Multienzyme Cascade on DNA Scaffold

Even though enzymatic fuel cells (EFCs) are considered as a promising green alternative method of power generation, much of their research has been limited to the use of a single enzyme as biocatalyst, leading to incomplete oxidation as well as limited range of simple fuels. In this work, enzymatic fuel cell utilizing multienzyme cascade system immobilized on DNA scaffold as anodic biocatalyst as well as high-surface area, electrically conductive carbonaceous nanofibers (CNFs) as electrodes was built for the first time to accommodate a complex molecule such as cellulose as fuel. Three cellulases and cellulose-binding domain (CBD) were expressed in E.Coli, purified using elastin-like polypeptide (ELP) and then conjugated with DNA linker using a fusion partner called HaloTag. All four enzymes and commercial glucose oxidase (GOx) were site-specifically immobilized on customized DNA template via hybridization for efficient cellulose hydrolysis and subsequent glucose oxidation. Varied number of enzymes were combined on the DNA scaffold and resulting reaction rate was compared to the same number of enzymes freely suspended in solution. Successful immobilization of enzymes on the DNA template increased the reaction rate up to 10-fold compared to when they were in solution, confirming the synergistic effect of the multiple enzymes immobilized on a scaffold. Investigation of temperature and pH effect revealed imitating the enzymes’ typical habitat (i.e. 37 oC and pH5) further increased the reaction rate of the multienzyme system. Carbonaceous nanofibers (CNFs) were used as the electrodes for both anode and cathode, which were first produced as polyacrylonitrile (PAN) nanofibers via electrospinning method. Three sets of design of experiments (DOEs) were used to systematically vary the solution, electrospinning, and environmental conditions. Analyses of the effect of the conditions on nanofiber diameter and bead density allowed for minimization of PAN nanofiber diameter down to 38 ± 7 nm with negligible bead density. PAN nanofibers were then converted to CNFs via a two-step heat treatment, and their resulting molecular structure confirmed by FT-IR and XRD was graphitic. Electrical conductivity characterized by 4-point probe measurement supported that CNFs were suitable for electrode applications, ranging from 1.2 to 7.4 S/cm. Electrochemical characterization of CNF mat electrode functionalized with GOx exhibited direct electron transfer from GOx cofactor to the CNF mat electrode at E=-0.63 V vs. Ag/AgCl. Anodic performance of CNF mat electrode functionalized with multienzyme system was electrochemically characterized by cyclic voltammetry, chronoamperometry and electrochemical impedance spectroscopy. Finally, cellulolytic enzymatic fuel cell was assembled with the multienzymatic CNF anode and CNF cathode, which was functionalized with bilirubin oxidase for oxygen reduction. Figure 1. Schematic representation of the cellulolytic enzymatic fuel cell with multienzyme cascade on DNA scaffold immobilized on carbonaceous nanofiber mat electrode. Components not drawn to scale. Figure 1

Performance evaluation of glucose oxidation reaction using biocatalysts adopting different quinone derivatives and their utilization in enzymatic biofuel cells

Glucose oxidase (GOx) and four different quinone derivatives (p-benzoquinone (BQ), naphthoquinone (NQ), anthraquinone (AQ) and 1,5-Dihydroxyanthraquinone (15DHAQ)) based biocomposites were embedded in polyethyleneimine (PEI) and then immobilized on carbon nanotube (CNT) substrate (CNT/PEI/Quinone/GOx). These catalysts were then used as the anodic biocatalysts for the enzymatic biofuel cell (EBC). According to the performance investigations of catalysts, the catalytic activity for glucose oxidation reaction (GOR) representing the electron transfer rate between GOx and glucose fuel is mostly enhanced in CNT/PEI/NQ/GOx. It is because two benzene rings of NQ play a role in attracting and releasing electrons effectively, increasing the catalytic activity for GOR, while other quinones have problems about attracting electrons (AQ and 15DHAQ) and wrong position of the reactive site for electron transfer (BQ). Excellent electron transfer rate constant (1.1 s-1) and Michaelis-Menten constant (0.99mM) are outstanding evidence for that. Furthermore, when the catalyst is utilized for EBC, high power density (57.4 μWcm-2) and high open circuit voltage (0.64 V) are accomplished.

Evaluation of Kefir as a New Anodic Biocatalyst Consortium for Microbial Fuel Cell.

Kefir, a combined consortium of bacteria and yeast encapsulated by a polymeric matrix of exopolysaccharides, was used as anodic biocatalyst in a two-chamber microbial fuel cell (MFC). Fermentation was followed during 72h and polarization curves were obtained from linear sweep voltammetry. The effect of methylene blue as charge-transfer mediator in the kefir metabolism was evaluated. UV/Vis spectrophotometry and cyclic voltammetry were applied to evaluate the redox state of the mediator and to characterize the electrochemical activity, whereas current interruption was used for internal resistance determination. Aiming to establish a relationship between the microbial development inside the anodic chamber with the generated power in the MFC, total titratable acidity, pH, viscosity, carbohydrate assimilation, and microbial counting were assayed. The kefir-based MFC demonstrated a maximum power density of 54mWm-2 after 24h fermentation, revealing the potential use of kefir as a biocatalyst for microbial fuel cells.

Enzymatic fuel cells with an oxygen resistant variant of pyranose-2-oxidase as anode biocatalyst

In enzymatic fuel cells (EnFCs), hydrogen peroxide formation is one of the main problems when enzymes, such as, glucose oxidase (GOx) is used due to the conversion of oxygen to hydrogen peroxide in the catalytic reaction. To address this problem, we here report the first demonstration of an EnFC using a variant of pyranose-2-oxidase (P2O-T169G) which has been shown to have low activity towards oxygen. A simple and biocompatible immobilisation approach incorporating multi-walled-carbon nanotubes within ferrocene (Fc)-Nafion film was implemented to construct EnFCs. Successful immobilisation of the enzymes was demonstrated showing 3.2 and 1.7-fold higher current than when P2O-T169G and GOx were used in solution, respectively. P2O-T169G showed 25% higher power output (maximum power density value of 8.45 ± 1.6 μW cm−2) and better stability than GOx in aerated glucose solutions. P2O-T169G maintained > 70% of its initial current whereas GOx lost activity > 90% during the first hour of 12 h operation at 0.15 V (vs Ag/Ag+). A different fuel cell configuration using gas-diffusion cathode and carbon paper electrodes were used to improve the power output of the fuel cell to 29.8 ± 6.1 µW cm−2. This study suggests that P2O-T169G with low oxygen activity could be a promising anode biocatalyst for EnFC applications.

Syntrophic association and performance of Clostridium, Desulfovibrio, Aeromonas and Tetrathiobacter as anodic biocatalysts for bioelectricity generation in dual chamber microbial fuel cell.

Anode chamber of a dual chamber microbial fuel cell (MFC) having raw landfill leachate was inoculated with consortium of sulphate-reducing bacteria (SRB) and sulphide-oxidizing bacteria (SOB) to study the phylogenetic architecture, function and mutualism of anolyte community developed in the reactor. Enriched microbial community was analysed with the help of Illumina MiSeq and indicated the dominance of Firmicutes (41.4%), Clostridia (36.4%) and Clostridium (12.9%) at phylum, class and genus level, respectively. Clostridium was associated with fermentation as well as transfer of electrons to the electrode mediated by ferredoxin. Desulfovibrio (6.7%), Aeromonas (6.6%) and Tetrathiobacter (9.8%) were SRB-SOB associated with direct electron transfer to the electrode. Community analysis disclosed a syntrophic association among novel Firmicutes and Proteobacteria species for bioelectricity generation and degradation of organic matter. Complete removal of chemical oxygen demand was observed from landfill leachate within 3days of inoculation. Lower oxidative slope and polarization resistance revealed from Tafel analysis backed the feasibility of electron transfer from microbes to anodic electrode and thus development of efficient anode-respiring community. Following enrichment and stabilization of the anodic community, maximum power density achieved was 9.15W/m3 and volumetric current density was 16.17A/m3. Simultaneous feeding with SRB-SOB and landfill leachate led to the enrichment of a novel, mutually interdependent microbial community capable of synchronized bioremediation of effluents rich in carbon, sulphate, nitrate and aromatic compounds.