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Galactose monitoring in individuals allows the prevention of harsh health conditions related to hereditary metabolic diseases like galactosemia. Current methods of galactose detection need development to obtain cheaper, more reliable, and more specific sensors. Enzyme-containing amperometric sensors based on galactose oxidase activity are a promising approach, which can be enhanced by means of their inclusion in a redox polymer coating. This strategy simultaneously allows the immobilization of the biocatalyst to the electroactive surface and hosts the electron shuttling units. An additional deposition of capping polymers prevents external interferences like ascorbic or uric acid as well as biofouling when measuring in physiological fuels. This work studies the protection effect of poly(2-methacryloyloxyethyl phosphorylcholine-co-glycidyl methacrylate (MPC) and polyvinylimidazole-polysulfostyrene (P(VI-SS)) when incorporated in the biosensor design for the detection of galactose in human plasma. [ABSTRACT FROM AUTHOR]

Photosynthetic hydrogen (photoH2) production is an elegant approach to storing solar energy. The most efficient strategy is to couple the hydrogen‐producing enzyme, the hydrogenase (H2ase), directly to photosystem I (PSI), which is a light‐driven nanomachine found in photosynthetic organisms. PSI–H2ase fusions have been tested in vivo and in vitro. Both approaches have each their specific advantages and drawbacks. Here, a system to combine both approaches by assembling PSI–H2ase fusions in vivo for in vitro photoH2 production is established. For this, cyanobacterial PSI–H2ase fusion mutants are generated and characterized concerning photoH2 production in vivo. The chimeric protein is purified and embedded in a redox polymer on an electrode where it successfully produces photoH2 in vitro. The combination of in vivo and in vitro processes comes along with reciprocal benefits. The in vivo assembly ensures that the chimeric protein is fully functional and suited for the fabrication of bioelectrodes in vitro. At the same time, the photoelectrochemical in vitro characterization now permits to analyze the assemblies in detail. This will open avenues to optimize in vivo and in vitro approaches for photoH2 production in a target‐oriented manner in the future. [ABSTRACT FROM AUTHOR]

Despite the success of continuous glucose measuring systems operating through the skin for about 14 days, long‐term implantable biosensors are facing challenges caused by the foreign‐body reaction. We present a conceptually new strategy using catalytic enzyme‐based biosensors based on a measuring sequence leading to minimum disturbance of the substrate equilibrium concentration by controlling the sensor between "on" and "off" state combined with short potentiometric data acquisition. It is required that the enzyme activity can be completely switched off and no parasitic side reactions allow substrate turnover. This is achieved by using an O2‐independent FAD‐dependent glucose dehydrogenase embedded within a crosslinked redox polymer. A short measuring interval allows the glucose concentration equilibrium to be restored quickly which enables the biosensor to operate under quasi‐equilibrium conditions. [ABSTRACT FROM AUTHOR]

An enzymatic biofuel cell is integrated on a screen‐printed electrode as a basis for a self‐powered biosensor. A glucose/O2 biofuel cell consisting of a pyrroloquinoline quinone‐dependent glucose dehydrogenase embedded within an Os‐complex modified redox polymer bioanode to oxidize glucose and a non‐limiting bilirubin oxidase‐based gas diffusion biocathode in the direct‐electron transfer regime for the reduction of O2 showed a glucose‐dependent current and power output. For full integration on a single screen‐printed electrode, a miniaturized agar salt bridge was introduced between the two bioelectrodes to ensure operation of the assembly in a two‐compartment configuration with each electrode operating at optimal conditions. [ABSTRACT FROM AUTHOR]

Using viologen‐based redox polymers to wire a variety of different hydrogenases to electrodes and gas diffusion electrodes is the basis to mitigate high potential deactivation of the enzyme, deactivation by molecular O2, as well as for high‐current density H2 oxidation bioanodes. To overcome electron transfer limitations by electron hopping within the viologen‐modified polymer film, a new redox polymer was designed with the highest possible viologen content together with monomers bearing crosslinking units. In combination with an immobilization sequence consisting of oxidative grafting of amino functions, covalent attachment of polymer units to these functionalities, and crosslinking of the polymer layers, an unprecedently fast electron transfer became possible. This enabled a very high current density normalized by the amount of the [NiFe] hydrogenase embedded within a viologen polymer on gas diffusion electrodes. [ABSTRACT FROM AUTHOR]

The use of enzymes as catalysts in chemical synthesis offers advantages in terms of clean and highly selective transformations. Galactose oxidase (GalOx) is a remarkable enzyme with several applications in industrial conversions as it catalyzes the oxidation of primary alcohols. We have investigated the wiring of GalOx with a redox polymer; this enables mediated electron transfer with the electrode surface for its potential application in biotechnological conversions. As a result of electrochemical regeneration of the catalytic center, the formation of harmful H2O2 is minimized during enzymatic catalysis. The introduced bioelectrode was applied to the conversion of bio‐renewable platform materials, with glycerol as model substrate. The biocatalytic transformations of glycerol and 5‐hydroxymethylfurfural (HMF) were investigated in a circular flow‐through setup to assess the possibility of substrate over‐oxidation, which is observed for glycerol oxidation but not during HMF conversion. [ABSTRACT FROM AUTHOR]

Photosystem I (PSI), a robust and abundant biomolecule capable of delivering high‐energy photoelectrons, has a great potential for the fabrication of light‐driven semi‐artificial bioelectrodes. Although possibilities have been explored in this regard, the true capabilities of this technology have not been achieved yet, particularly for their use as bioanodes. Here, the use of PSI Langmuir monolayers and their electrical wiring with specifically designed redox polymers is shown, ensuring an efficient mediated electron transfer as the basis for the fabrication of an advanced biophotoanode. The bioelectrode is rationally implemented and optimized for enabling the generation of substantial photocurrents of up to 17.6 µA cm−2 and is even capable of delivering photocurrents at potentials as low as −300 mV vs standard hydrogen electrode, surpassing the performance of comparable devices. To highlight the applicability of the developed light‐driven bioanode, a biophotovoltaic cell is assembled in combination with a gas‐breathing biocathode. The assembly operates in a single compartment cell and delivers considerable power outputs at large cell voltages. The implemented biophotoanode constitutes an important step toward the development of advanced biophotovoltaic devices. [ABSTRACT FROM AUTHOR]

Redox polymers with distinct redox units have been long recognized for their pseudocapacitive and reversible charge storage behaviour. Many systems investigated so far have utilized organic electrolytes and/or have coupled a redox polymer half-cell to a non-polymer counter electrode. However, due to safety and sustainability considerations, aqueous electrolyte based charge storage in all-polymer configurations is considered a promising option for possible future applications. We present a strategy based on pseudocapacitive charge storage in Osmium-complex and viologen-modified redox polymers with specifically designed poly(vinylimidazole)- and poly(vinylpyridine)-based backbones. We couple both redox polymers in an aqueous battery configuration, demonstrating Nernst-potential driven energy storage. Electrochemical characterization in a concentric three-electrode Swagelok cell and coin cells reveals stable reversible capacities over more than 1800 cycles, with nearly quantitative coulombic efficiencies (>99.4%) for the coin cells. [ABSTRACT FROM AUTHOR]

We report on a photobioelectrochemical fuel cell consisting of a glucose‐oxidase‐modified BiFeO3 photobiocathode and a quantum‐dot‐sensitized inverse opal TiO2 photobioanode linked to FAD glucose dehydrogenase via a redox polymer. Both photobioelectrodes are driven by enzymatic glucose conversion. Whereas the photobioanode can collect electrons from sugar oxidation at rather low potential, the photobiocathode shows reduction currents at rather high potential. The electrodes can be arranged in a sandwich‐like manner due to the semi‐transparent nature of BiFeO3, which also guarantees a simultaneous excitation of the photobioanode when illuminated via the cathode side. This tandem cell can generate electricity under illumination and in the presence of glucose and provides an exceptionally high OCV of about 1 V. The developed semi‐artificial system has significant implications for the integration of biocatalysts in photoactive entities for bioenergetic purposes, and it opens up a new path toward generation of electricity from sunlight and (bio)fuels. [ABSTRACT FROM AUTHOR]

Well‐defined assemblies of photosynthetic protein complexes are required for an optimal performance of semi‐artificial energy conversion devices, capable of providing unidirectional electron flow when light‐harvesting proteins are interfaced with electrode surfaces. We present mixed photosystem I (PSI) monolayers constituted of native cyanobacterial PSI trimers in combination with isolated PSI monomers from the same organism. The resulting compact arrangement ensures a high density of photoactive protein complexes per unit area, providing the basis to effectively minimize short‐circuiting processes that typically limit the performance of PSI‐based bioelectrodes. The PSI film is further interfaced with redox polymers for optimal electron transfer, enabling highly efficient light‐induced photocurrent generation. Coupling of the photocathode with a [NiFeSe]‐hydrogenase confirms the possibility to realize light‐induced H2 evolution. [ABSTRACT FROM AUTHOR]

We report an organic redox‐polymer‐based electroenzymatic nitrogen fixation system using a metal‐free redox polymer, namely neutral‐red‐modified poly(glycidyl methacrylate‐co‐methylmethacrylate‐co‐poly(ethyleneglycol)methacrylate) with a low redox potential of −0.58 V vs. SCE. The stable and efficient electric wiring of nitrogenase within the redox polymer matrix enables mediated bioelectrocatalysis of N3−, NO2− and N2 to NH3 catalyzed by the MoFe protein via the polymer‐bound redox moieties distributed in the polymer matrix in the absence of the Fe protein. Bulk bioelectrosynthetic experiments produced 209±30 nmol NH3 nmol MoFe−1 h−1 from N2 reduction. 15N2 labeling experiments and NMR analysis were performed to confirm biosynthetic N2 reduction to NH3. [ABSTRACT FROM AUTHOR]

The incorporation of highly active but also highly sensitive catalysts (e.g. the [FeFe] hydrogenase from Desulfovibrio desulfuricans) in biofuel cells is still one of the major challenges in sustainable energy conversion. We report the fabrication of a dual‐gas diffusion electrode H2/O2 biofuel cell equipped with a [FeFe] hydrogenase/redox polymer‐based high‐current‐density H2‐oxidation bioanode. The bioanodes show benchmark current densities of around 14 mA cm−2 and the corresponding fuel cell tests exhibit a benchmark for a hydrogenase/redox polymer‐based biofuel cell with outstanding power densities of 5.4 mW cm−2 at 0.7 V cell voltage. Furthermore, the highly sensitive [FeFe] hydrogenase is protected against oxygen damage by the redox polymer and can function under 5 % O2. [ABSTRACT FROM AUTHOR]

Variants of the highly active [NiFeSe] hydrogenase from D. vulgaris Hildenborough that exhibit enhanced O2 tolerance were used as H2‐oxidation catalysts in H2/O2 biofuel cells. Two [NiFeSe] variants were electrically wired by means of low‐potential viologen‐modified redox polymers and evaluated with respect to H2‐oxidation and stability against O2 in the immobilized state. The two variants showed maximum current densities of (450±84) μA cm−2 for G491A and (476±172) μA cm−2 for variant G941S on glassy carbon electrodes and a higher O2 tolerance than the wild type. In addition, the polymer protected the enzyme from O2 damage and high‐potential inactivation, establishing a triple protection for the bioanode. The use of gas‐diffusion bioanodes provided current densities for H2‐oxidation of up to 6.3 mA cm−2. Combination of the gas‐diffusion bioanode with a bilirubin oxidase‐based gas‐diffusion O2‐reducing biocathode in a membrane‐free biofuel cell under anode‐limiting conditions showed unprecedented benchmark power densities of 4.4 mW cm−2 at 0.7 V and an open‐circuit voltage of 1.14 V even at moderate catalyst loadings, outperforming the previously reported system obtained with the [NiFeSe] wild type and the [NiFe] hydrogenase from D. vulgaris Miyazaki F. [ABSTRACT FROM AUTHOR]

The fabrication and electrochemical evaluation of transparent photoelectrodes consisting of Photosystem I (PSI) or Photosystem II (PSII) is described, which are embedded and electrically wired by a redox polymer. The fabrication process is performed by an automated airbrush-type spray coating system, which ensures controlled and scalable electrode preparation. As proof of concept, electrodes with a surface area of up to 25 cm2 were prepared. The macro-porous structure of the indium tin oxide electrodes allows a high loading of the photoactive protein complexes leading to enhanced photocurrents, which are essential for potentially technologically relevant solar-powered devices. In addition, we show that unpurified crude PSII extracts, which can be provided in comparatively high yields for electrode modification, are suitable for photoelectrode fabrication with comparable photocurrent densities. [ABSTRACT FROM AUTHOR]

We report the fabrication of a polymer/enzyme‐based biosupercapacitor (BSC)/biofuel cell (BFC) hybrid device with an optimized cell voltage that can be switched on demand from energy conversion to energy storage mode. The redox polymer matrices used for the immobilization of the biocatalyst at the bioanode and biocathode act simultaneously as electron relays between the integrated redox enzymes and the electrode surface (BFC) and as pseudocapacitive charge storing elements (BSC). Moreover, owing to the self‐charging effect based on the continuously proceeding enzymatic reaction, a Nernstian shift in the pseudocapacitive elements, that is, in the redox polymers, at the individual bioelectrodes leads to a maximized open circuit voltage of the device in both operating modes. Comparison with a conventional fuel cell design, that is, using redox mediators with redox potentials that are close to the potentials of the used redox proteins, indicates that the novel hybrid device shows a similar voltage output. Moreover, our results demonstrate that the conventional design criteria commonly used for the development of redox polymers for the use in biofuel cells have to be extended by considering the effect of a Nernstian shift towards the potentials of the used biocatalysts in those pseudocapacitive elements. [ABSTRACT FROM AUTHOR]

We report a prototype air‐breathing carbon cloth‐based electrode that was fabricated starting from a commercially available screen‐printed electrode equipped with a transparent ITO working electrode (DropSens, ref. ITO10). The fabrication of the air‐breathing electrodes is straightforward, shows satisfactory reproducibility and a good electrochemical response as evaluated by means of [Fe(CN)6]3−/4− voltammetry. The gas‐diffusion electrodes were successfully modified with the O2 reducing enzyme bilirubin oxidase from Myrothecium verrucaria in a direct electron transfer regime. The enzyme modified electrodes showed a remarkable high current density for O2 reduction in passive air‐breathing mode of up to 5 mA cm−2. Moreover, the enzyme modified electrodes were applied as O2 reducing biocathodes in a glucose/air enzymatic biofuel cell in combination with a high current density glucose oxidase/redox polymer bioanode. The biofuel cell provides a high maximum power density of (0.34±0.02) mW cm−2 at 0.25 V. The straightforward design, low cost and the high reproducibility of these electrodes are considered as basis for standardized measurements under gas‐breathing conditions and for high throughput screening of gas converting (bio‐)catalysts. [ABSTRACT FROM AUTHOR]

A biohybrid photobioanode mimicking the Z‐scheme has been developed by functional integration of photosystem II (PSII) and PbS quantum dots (QDs) within an inverse opal TiO2 architecture giving rise to a rather negative water oxidation potential of about −0.55 V vs. Ag/AgCl, 1 m KCl at neutral pH. The electrical linkage between both light‐sensitive entities has been established through an Os‐complex‐modified redox polymer (POs), which allows the formation of a multi‐step electron‐transfer chain under illumination starting with the photo‐activated water oxidation at PSII followed by an electron transfer from PSII through POs to the photo‐excited QDs and finally to the TiO2 electrode. The photobioanode was coupled to a novel, transparent, inverse‐opal ATO cathode modified with an O2‐reducing bilirubin oxidase for the construction of a H2O/O2 photobioelectrochemical cell reaching a high open‐circuit voltage of about 1 V under illumination. A Z‐scheme mimic based on photosystem II wired to a quantum‐dot‐modified inverse opal TiO2 electrode allows H2O oxidation at very low potential. The sandwich‐like arrangement of this photobioanode and a transparent ATO electrode hosting bilirubin oxidase are used for the construction of a H2O/O2 photobioelectrochemical cell, which can be illuminated through the transparent biocathode. [ABSTRACT FROM AUTHOR]

Abstract: In the development of biofuel cells great effort is dedicated to achieving outstanding figures of merit, such as high stability, maximum power output, and a large open circuit voltage. Biofuel cells with immobilized redox mediators, such as redox polymers with integrated enzymes, show experimentally a substantially higher open circuit voltage than the thermodynamically expected value. Although this phenomenon is widely reported in the literature, there is no comprehensive understanding of the potential shift, the high open circuit voltages have not been discussed in detail, and hence they are only accepted as an inherent property of the investigated systems. We demonstrate that this effect is the result of a Nernstian shift of the electrode potential when catalytic conversion takes place in the absence or at very low current flow. Experimental evidence confirms that the immobilization of redox centers on the electrode surface results in the assembled biofuel cell delivering a higher power output because of charge storage upon catalytic conversion. Our findings have direct implications for the design and evaluation of (bio)fuel cells with pseudocapacitive elements. [ABSTRACT FROM AUTHOR]

Abstract: The development of O2 tolerant glucose sensors based on the highly active and robust enzyme glucose oxidase is still a major challenge because of the competition between the natural electron acceptor O2 and free‐diffusing or polymer‐bound artificial electron acceptors. We report the fabrication of a glucose oxidase based bioanode that operates under ambient conditions. Combination of this bioanode with a bilirubin oxidase based biocathode enabled the fabrication of a glucose/O2 powered biofuel cell as integrated power source for a self‐powered device. Glucose oxidase at the anode was electrically wired via a low‐potential redox polymer, i. e. a Toluidine Blue‐modified poly(methacrylate) based polymer, that ensures a high open‐circuit voltage of the biofuel cell but also catalytically reduces O2 and hence requires a protection shield for measurements under ambient conditions. The sensing layer was deposited by means of potential pulse‐assisted co‐deposition of glucose oxidase within the redox polymer and was protected from O2 by a newly proposed lactate oxidase/catalase based O2 removal layer that was immobilized within a hydrophilic redox‐silent polymer on top of the sensing layer. The protection layer was powered by lactate, a natural component in human blood. The biofuel cell exhibited an OCV of ca. 650 mV and the power output was dependent on the glucose concentration without any interference from oxygen providing that lactate was available in the analyte solution. [ABSTRACT FROM AUTHOR]

The Ni(P2N2)2 catalysts are among the most efficient non-noble-metal based molecular catalysts for H2 cycling. However, these catalysts are O2 sensitive and lack long term stability under operating conditions. Here, we show that in a redox silent polymer matrix the catalyst is dispersed into two functionally different reaction layers. Close to the electrode surface is the “active” layer where the catalyst oxidizes H2 and exchanges electrons with the electrode generating a current. At the outer film boundary, insulation of the catalyst from the electrode forms a “protection” layer in which H2 is used by the catalyst to convert O2 to H2O, thereby providing the “active” layer with a barrier against O2. This simple but efficient polymer-based electrode design solves one of the biggest limitations of these otherwise very efficient catalysts enhancing its stability for catalytic H2 oxidation as well as O2 tolerance. [ABSTRACT FROM AUTHOR]

An intrinsic self-charging biosupercapacitor built on a unique concept for the fabrication of biodevices based on redox polymers is presented. The biosupercapacitor consists of a high-potential redox polymer based bioanode and a low-potential redox polymer based biocathode in which the potentials of the electrodes in the discharged state show an apparent potential mismatch Eanode>Ecathode and prevent the use of the device as a conventional biofuel cell. Upon charging, the potentials of the electrodes are shifted to more positive (cathode) and more negative (anode) values because of a change in the aox-to-aredratio within the redox polymer matrix. Hence, a potential inversion occurs in the charged state (Eanodecathode) and an open circuit voltage of >0.4 V is achieved and the biodevice acts as a true biosupercapacitor. The bioanode consists of a novel specifically designed high-potential Os complex modified polymer for the efficient immobilization and electrical wiring of glucose converting enzymes, such as glucose oxidase and flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase. The cathodic side is constructed from a low-potential Os complex modified polymer integrating the O2 reducing enzyme, bilirubin oxidase. The large potential differences between the redox polymers and the prosthetic groups of the biocatalysts ensure fast and efficient charging of the biodevice. [ABSTRACT FROM AUTHOR]

We describe the fabrication of a self-powered ethanol biosensor comprising a β-NAD+-dependent alcohol dehydrogenase (ADH) bioanode and a bienzymatic alcohol oxidase (AOx) and horseradish peroxidase (HRP) biocathode. β-NAD+ is regenerated by means of a specifically designed phenothiazine dye (i. e. toluidine blue, TB) modified redox polymer in which TB was covalently anchored to a hexanoic acid tethered poly(4-vinylpyridine) backbone. The redox polymer acts as an immobilization matrix for ADH. Using a carefully chosen anchoring strategy through the formation of amide bonds, the potential of the TB-based mediator is shifted to more positive potentials, thus preventing undesired O2 reduction. To counterbalance the rather high potential of the TB-modified polymer, and thus the bioanode, a high-potential AOx/HRP-based biocathode is suggested. HRP is immobilized in a direct-electron-transfer regime on screen-printed graphite electrodes functionalized with multi-walled carbon nanotubes. The nanostructured cathode ensures the wiring of the iron-oxo complex within oxidized HRP, and thus a high potential for the reduction of H2O2 of about +550 mV versus Ag/AgCl/3 M KCl. The proposed biofuel cell exhibits an open-circuit voltage (OCV) of approximately 660 mV and was used as self-powered device for the determination of the ethanol content in liquor. [ABSTRACT FROM AUTHOR]

We propose the very first 'Nernstian biosupercapacitor', a biodevice based on only one redox polymer: poly(vinyl imidazole- co-allylamine)[Os(bpy)2Cl], and two biocatalysts. At the bioanode PQQ-dependent glucose dehydrogenase reduces the Os3+ moieties at the polymer to Os2+ shifting the Nernst potential of the Os3+/Os2+ redox couple to negative values. Concomitantly, at the biocathode the reduction of O2 by means of bilirubin oxidase embedded in the same redox polymer leads to the oxidation of Os2+ to Os3+ shifting the Nernst potential to higher values. Despite the use of just one redox polymer an open circuit voltage of more than 0.45 V was obtained during charging and the charge is stored in the redox polymer at both the bioanode and the biocathode. By connecting both electrodes via a predefined resistor a high power density is obtained for a short time exceeding the steady state power of a corresponding biofuel cell by a factor of 8. [ABSTRACT FROM AUTHOR]

Benzoxazine oligomers synthesized by a Mannich type reaction of bisphenol A, tetraethylenepentamine and formaldehyde were electrochemically crosslinked in presence of both an Os-complex modified poly(methacrylate) polymer and glucose oxidase. The crosslinking led to the formation of a biocatalytically active layer on an electrode surface exhibiting a swelling process after immersion in an electrolyte solution containing glucose most likely due to the local decrease of the pH value upon glucose oxidation. Optimization of the poly(benzoxazine) to Os-complex modified poly(methacrylate) ratio was performed leading to a reagentless glucose biosensor with improved stability. [ABSTRACT FROM AUTHOR]

Benzoxazine-based redox polymers bearing Os complexes are synthesized and used as an immobilization matrix for glucose oxidase (GOx) as a model system for a reagentless biosensor. The polymers are formed by electrochemically induced anodic polymerization of the corresponding benzoxazine monomers modified with Os complexes. The precursors are synthesized in a Mannich-type reaction between bisphenol A, formaldehyde, and the corresponding Os complexes or ligands, which contain free amino groups. The Os complexes are redox active within the polymer films, and thus, can be used as redox relays for the electron transfer between the electrode surface and the prosthetic group within the enzyme. Entrapment of GOx within the poly(benzoxazine) film is achieved successfully, as shown by the biocatalytic activity of the poly(benzoxazine)/ GOx films upon the addition of glucose. [ABSTRACT FROM AUTHOR]

A variety of Os-complex modified redox polymers were synthesized and optimized for an improved bioelectrochemical communication with PQQ-sGDH. The properties of the polymers were varied by changing the monomers composition leading to improved immobilization and increased enzyme loading. Three Os-complexes were designed exhibiting different ligands but similar redox potentials adapted for efficient electron transfer with PQQ-sGDH. Effects of the ligand sphere, polymer backbone and length of the tether chain between Os-complex and polymer backbone as well as the impact of different bifunctional crosslinkers on the catalytic current were evaluated. The optimized bioelectrodes exhibited significantly improved catalytic currents and stability. [ABSTRACT FROM AUTHOR]

The integration of photosystem 1 in redox hydrogels based on Os-complexes modified redox polymers on electrodes yields efficient photocathodes. The generation of high photocurrent relies on high loading in PS 1 and fast electron transfer rates from the electrode to PS 1. The interaction between the redox polymer and PS 1 influences both the loading in protein and the electron transfer rates. Since PS1 exhibits extended hydrophobic regions, polymers with similar properties may favor attractive interactions. Here we investigate three approaches to confer hydrophobicity to the redox polymer. We demonstrate that the pyridine functionality enables to switch, via basic pH values, the polymer properties from hydrophilic to hydrophobic. The transition triggers a hydrogel collapse which allows for efficient entrapment of PS 1. In addition the hydrophobic-hydrophilic balance was tuned by the addition of hydrophobic group in i) the polymer backbone and ii) as substituents at the Os-complex. The increased hydrophobicity of the backbone results in higher photocurrents from PS 1 integrated in the corresponding hydrogel. On the other hand, further increasing hydrophobicity of the redox relay decreases the photocurrent due to either lower mobility of the Os-complexes or poor interaction with the hydrophilic site where the redox center of PS1 is located. [ABSTRACT FROM AUTHOR]

Bio-electrocatalysis, galactose oxidase, glycerol oxidation, modified electrodes, redox polymers Keywords: bio-electrocatalysis; galactose oxidase; glycerol oxidation; modified electrodes; redox polymers EN bio-electrocatalysis galactose oxidase glycerol oxidation modified electrodes redox polymers 1 1 1 05/27/22 20220525 NES 220525 B An enzymatic bioelectrode b made up of galactose oxidase electrically wired with a redox polymer was used for the electrocatalytic oxidation of bio-renewable substrates with minimal influence from ambient O SB 2 sb . [Extracted from the article]

A miniaturized pH sensor based on the voltammetric determination of the potential difference between a pH dependent toluidine blue O modified redox polymer and a pH independent osmium complex modified redox polymer was developed. Gold micro electrodes of various dimensions were modified with the two redox polymers by non-manual deposition procedures. Square wave voltammetry was used to determine the oxidation potentials of the polymer bound redox species allowing the precise determination of the pH value of the electrolyte solution without the need of a stable reference electrode. [ABSTRACT FROM AUTHOR]

Carbon nanotubes (CNTs) grown on carbon cloth substantially increased the surface area of the electrodes. Carbon cloths were pretreated with HNO3 vapor before CNTs growth and electrochemically oxidized afterwards. The CNT-modified carbon cloths were characterized using scanning electron microscopy, Raman spectroscopy, cyclic voltammetry and electrochemical impedance spectroscopy. Biofuel cells based on these CNT-modified electrode materials using Laccase from Trametes hirsuta and cellobiose dehydrogenase from Myriococcum thermophilium entrapped in specifically designed Os-complex modified redox polymers showed a power density of 5.87 µW/cm2 which is 125 fold enhanced as compared with electrodes prepared on untreated carbon cloth. [ABSTRACT FROM AUTHOR]

We report on the development of a bi-layer bi-enzyme biosensor architecture using different peroxidases and alcohol oxidase from Hansenula polymorpha C-105 as biological recognition elements. The sensor architecture comprises a first layer containing either horseradish peroxidase or royal palm tree peroxidase crosslinked with an Osmium complex-modified redox hydrogel. On top, a second layer was formed by electrochemically induced precipitation of a cathodic electrodeposition paint simultaneously entrapping alcohol oxidase isolated from a genetically modified strain of Hansenula polymorpha C-105. The sensor architecture was optimized with respect to effective electron transfer and stability of the enzyme. The main characteristics of the biosensors are an apparent maximal current Imax app of 572–940 nA, an apparent Michaelis constant KM app of 9.5 mM, a sensitivity of 60–98 nA mM−1 and an improved operational stability represented by a deactivation constant of 1.5–2.0 × 10−4min−1. [ABSTRACT FROM AUTHOR]

If the analyte does not only change the electrochemical but also the optical properties of the electrode/solution interface, the spatial resolution of an electrochemical sensor can be substantially enhanced by combining the electrochemical sensor with optical microscopy. In order to demonstrate this, electrochemical biosensors for the detection of hydrogen peroxide and glucose were developed by drop casting enzyme and redox polymer mixtures onto planar, optically transparent electrodes. These biosensors generate current signals proportional to the analyte concentration via a reaction sequence which ultimately changes the oxidation state of the redox polymer. Images of the interface of these biosensors were acquired using bright field reflected light microscopy (BFRLM). Analysis showed that the intensity of these images is higher when the redox polymer is oxidized than when it is reduced. It also revealed that the time needed for the redox polymer to change oxidation state can be assayed optically and is dependent on the concentration of the analyte. By combining the biosensor for hydrogen peroxide detection with BFRLM, it was possible to determine hydrogen peroxide in concentrations as low as 12.5 µM with a spatial resolution of 12 µm × 12 µm, without the need for the fabrication of microelectrodes of these dimensions. [ABSTRACT FROM AUTHOR]

Metal-free organic redox mediators, whose redox potentials can be tuned according to the intended enzyme system and application, are emerging as alternative mediators for the design of improved enzyme-based biosensors and biofuel cells. We report a novel class of metal-free redox polymer containing a polymer-tethered dicyano viologen mediator for the immobilization of FAD-dependent glucose dehydrogenase (FAD-GDH) on graphite electrodes. Incorporation of strong electron-withdrawing cyano groups to the viologen moiety ensures the formation of high potential viologen-based redox polymer with an envisaged pH-independent redox potential close to 50 mV (vs Ag /AgCl/ 3 M KCl) and a negligible impact on oxygen reduction. The bioelectrode comprising of newly designed redox polymer and FAD-GDH showed enhanced electrocatalytic activity in the presence of glucose, confirming effective wiring of FAD-GDH, if embedded within the redox polymer. [Display omitted] • A novel viologen-based redox polymer with high redox potential was designed. • The redox polymer shows negligible activity for the oxygen reduction reaction. • The redox polymer exhibits a low potential to prevent oxidation of interferents. • The redox polymer efficiently wires FAD-GDH for enzymatic biosensors. • The redox polymer contains crosslinking sites for stable immobilization. [ABSTRACT FROM AUTHOR]

Monitoring galactose levels in dairy products can help to prevent severe complications with a hereditary metabolic disease such as galactosemia, a life-threatening disease. The current state of the art requires the development of less expensive, more reliable and specific methods to determine galactose levels in food in a practical way. We report the development and optimization of an amperometric biosensor for determination of galactose in dairy products based on galactose oxidase (GaOx) co-immobilized with an osmium-complex modified redox polymer on glassy carbon electrodes. To attain the maximum catalytic currents based on mediated electron transfer, two Os-complex based polymers with different redox potentials and different enzyme:redox polymer ratios were studied. The optimized GaOx-modified electrode gave a maximum electrocatalytic response of galactose oxidation that was not affected by the presence of O 2 , indicating fast wiring of the enzyme by the Os-complex modified redox polymer. The biosensor that gave the best analytical parameters for galactose detection was further tested for measuring galactose concentration in lactose-containing and lactose-free milk and yogurt samples under aerobic conditions. The results obtained with the amperometric biosensor were validated by high-performance anion-exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD). [ABSTRACT FROM AUTHOR]

Following inspiration by natural photosynthesis, the design and fabrication of semi-artificial biophotoelectrochemical devices able to harvest solar energy and aiming on the implementation of green and sustainable energy conversion systems is presently an important field of research. Here we present the development of a fully light-driven biosupercapacitor fabricated by incorporation of isolated photosystem 2 and photosystem 1 protein complexes embedded within the same Os-complex modified redox polymer. By this, light energy is stored at both electrodes within the polymer-based pseudocapacitive matrix in the form of Os3+ centers at the photosystem1-based biocathode and in the form of Os2+ centers at the photosystem 2-based bioanode. The stored energy can be released on demand into bursts of electricity. Due to the purely light-driven self-charging process, the biosupercapacitor provided a power output of 1.0 μW cm−2 after 200 s charging time. Moreover, the use of different electrode materials and their implication on the performance of the implemented biodevice is evaluated. Image 1 [ABSTRACT FROM AUTHOR]

Abstract In the development of semi-artificial biophotovoltaic assemblies, deeper understanding of electrochemical processes is required to achieve functional and efficient devices. Evaluation of photosystem 2 embedded in an Os-complex modified redox polymer using scanning photoelectrochemical microscopy (SPECM) provides insight into the intricate electrochemical processes of the immobilized protein complex and its electrical communication pathways with the redox tethers of the polymer matrix. The use of local irradiation during an SPECM array scan prevents sample inactivation prior to analysis. Moreover, the simultaneously possible collection of partially reduced oxygen species in the form of hydrogen peroxide confirms the presence of competing charge transfer pathways involved in the reduction of oxygen at the chlorophyll pigments upon irradiation of the sample. In addition, evaluation of photocurrent in the presence of an inhibitor that blocks the terminal plastoquinone Q B binding site of the photosystem reveals electrochemical communication between the intermediate plastoquinone Q A and the redox polymer. The obtained information proves to be relevant for further design and optimization of devices for technological applications. Graphical abstract Image 1 [ABSTRACT FROM AUTHOR]

Despite the availability of numerous electroanalytical methods for phosphate quantification, practical implementation in point-of-use sensing remains virtually nonexistent because of interferences from sample matrices or from atmospheric O 2 . In this work, phosphate determination is achieved by the purine nucleoside phosphorylase (PNP) catalyzed reaction of inosine and phosphate to produce hypoxanthine which is subsequently oxidized by xanthine oxidase (XOx), first to xanthine and then to uric acid. Both PNP and XOx are integrated in a redox active Os-complex modified polymer, which not only acts as supporting matrix for the bienzymatic system but also shuttles electrons from the hypoxanthine oxidation reaction to the electrode. The bienzymatic cascade in this second generation phosphate biosensor selectively delivers four electrons for each phosphate molecule present. We introduced an additional electrochemical process involving uric acid oxidation at the underlying electrode. This further enhances the anodic current (signal amplification) by two additional electrons per analyte molecule which mitigates the influence of electrochemical interferences from the sample matrix. Moreover, while the XOx catalyzed reaction is sensitive to O 2 , the uric acid production and therefore the delivery of electrons through the subsequent electrochemical process are independent of the presence of O 2 . Consequently, the electrochemical process counterbalances the O 2 interferences, especially at low phosphate concentrations. Importantly, the electrochemical uric acid oxidation specifically reports on phosphate concentration since it originates from the product of the bienzymatic reactions. These advantageous properties make this bioelectrochemical-electrochemical cascade particularly promising for point-of-use phosphate measurements. [ABSTRACT FROM AUTHOR]

We report the fabrication of an amperometric NADH biosensor system that employs an allosterically modulated bacterial reductase in an adapted osmium(III)-complex-modified redox polymer film for analyte quantification. Chains of complexed Os(III) centers along matrix polymer strings make electrical connection between the immobilized redox protein and a graphite electrode disc, transducing enzymatic oxidation of NADH into a biosensor current. Sustainable anodic signaling required (1) a redox polymer with a formal potential that matched the redox switch of the embedded reductase and avoided interfering redox interactions and (2) formation of a cross-linked enzyme/polymer film for stable biocatalyst entrapment. The activity of the chosen reductase is enhanced upon binding of an effector, i.e. p-hydroxy-phenylacetic acid (p-HPA), allowing the acceleration of the substrate conversion rate on the sensor surface by in situ addition or preincubation with p-HPA. Acceleration of NADH oxidation amplified the response of the biosensor, with a 1.5-fold increase in the sensitivity of analyte detection, compared to operation without the allosteric modulator. Repetitive quantitative testing of solutions of known NADH concentration verified the performance in terms of reliability and analyte recovery. We herewith established the use of allosteric enzyme modulation and redox polymer-based enzyme electrode wiring for substrate biosensing, a concept that may be applicable to other allosteric enzymes. [ABSTRACT FROM AUTHOR]

• A glucose oxygen biofuel cell/glucose sensor with a non-limiting biocathode. • Three types of bilirubin oxidase were evaluated in direct and mediated electron transfer. • The bilirubin oxidase from Magnaporthe oryzae (Mo- BOD) showed the highest oxygen reduction current. A new redox polymer/bilirubin oxidase (BOD)-based gas diffusion electrode was designed to be implemented as the non-current and non-stability limiting biocathode in a glucose/O 2 biofuel cell that acts as a self-powered glucose biosensor. For the proof-of-concept, a bioanode comprising the Os-complex modified redox polymer P(VI- co -AA)-[Os(bpy) 2 Cl]Cl and FAD-dependent glucose dehydrogenase to oxidize the analyte was used. In order to develop an optimal O 2 -reducing biocathode for the biofuel cell Mv -BOD as well as Bp -BOD and Mo -BOD have been tested in gas diffusion electrodes in direct electron transfer as well as in mediated electron transfer immobilized in the Os-complex modified redox polymer P(VI- co -AA)-[Os(diCl-bpy) 2 ]Cl 2. The resulting biofuel cell exhibits a glucose-dependent current and power output in the concentration region between 1 and 10 mM. To create a more realistic test environment, the performance and long-term stability of the biofuel cell-based self-powered glucose biosensor has been investigated in a flow-through cell design. [ABSTRACT FROM AUTHOR]

[Display omitted] • A bioelectrode constituted by a folded assembly of PSI monolayers is introduced. • The influence of incident light angle on photocurrent generation is investigated. • The folded assembly is shown to provide an improved utilization of incident light. In the fabrication of photosystem I (PSI)-based biodevices, the use of multilayered architectures aims to maximize the absorption of incident light that can be converted into high-energy electrons. The challenge in this strategy is to overcome the large driving force imposed by the photoinduced potential difference between the two terminal redox centers that are located at opposite sides of PSI, which translates into charge recombination resulting in sub-optimal performance of commonly implemented systems. The integration of PSI monolayers with electrodes using the Langmuir-Blodgett technique enables a preferential anisotropic orientation of PSI in a tightly packed structure, which minimizes short-circuiting processes and aids to improve the performance of PSI-based biodevices. However, the practical application of PSI monolayer-based biodevices is limited due to the small loading of immobilized PSI molecules, leading to overall low utilization of incident light. Inspired by the stacked arrangements of thylakoids in nature, we demonstrate the fabrication of biomimetic structures using multiple PSI monolayers assembled into a folded architecture to improve light absorption and with that the performance of the overall photoelectrode. [ABSTRACT FROM AUTHOR]

[Display omitted] • Boronic acid modification of C. vulgaris improves the crosslinking with redox polymer. • The bioanode's action spectrum confirms the photosystems as electron source. • Scanning photoelectrochemical microscopy suggests O 2 consumption upon illumination. • The bioanode is coupled to a bilirubin oxidase biocathode for a proof-of-concept biophotovoltaic cell. Green microalgae are gaining attention in the renewable energy field due to their ability to convert light into energy in biophotovoltaic (BPV) cells. The poor exogenous electron transfer kinetics of such microorganisms requires the use of redox mediators to improve the performance of related biodevices. Redox polymers are advantageous in the development of subcellular-based BPV devices by providing an improved electron transfer while simultaneously serving as immobilization matrix. However, these surface-confined redox mediators have been rarely used in microorganism-based BPVs. Since electron transfer relies on the proximity between cells and the redox centres at the polymer matrix, the development of molecularly tailored surfaces is of great significance to fabricate more efficient BPV cells. We propose a bioanode integrating Chlorella vulgaris embedded in an Os complex-modified redox polymer. Chlorella vulgaris cells are functionalized with 3-aminophenylboronic acid that exhibits high affinity to saccharides in the cell wall as a basis for an improved integration with the redox polymer. Maximum photocurrents of (5 ± 1) µA cm−2 are achieved. The developed bioanode is further coupled to a bilirubin oxidase-based biocathode for a proof-of-concept BPV cell. The obtained results encourage the optimization of electron-transfer pathways toward the development of advanced microalgae-based biophotovoltaic devices. [ABSTRACT FROM AUTHOR]

Abstract: The design of biofuel cell anodes with substantially decreased potential is a prerequisite for the development of biofuel cells with large open-circuit voltage and power density. Redox polymers with covalently attached phenothiazine derivatives such of thionine acetate, toluidine blue, azure B simultaneously providing epoxide functions for covalent binding to suitably modified electrode surfaces and crosslinking were synthesized and evaluated for their ability to transfer electrons from the FAD cofactor of the flavodehydrogenase domain of cellobiose dehydrogenase from Myriococcum thermophilum (FAD-MtCDH), the flavodehydrogenase domain of cellobiose dehydrogenase from Corynascus thermophilus (FAD-CtCDH), or glucose oxidase from Aspergillus niger (GOx). Polymer/enzyme films were covalently bound via polymer bound epoxy groups to terminal amino functions introduced to graphite electrode surfaces by electrochemically induced grafting of diaminoheptane or Boc-protected ethylene diamine (EDA). The electrodes were optimized for biocatalytic glucose oxidation with respect to the hydrophilicity of the polymer backbone, the nature of the phenothiazine derivative, the pH value, as well as the relative amount of enzyme, polymer and crosslinker. Biofuel cells based on toluidine blue-modified redox polymers with integrated FAD-MtCDH, FAD-CtCDH, or GOx in combination with a bilirubin oxidase based biocathode exhibited open-circuit voltages of more than 0.7V and maximum power densities in the range of 4 to 6μWcm−2 at a pH value of 7.8. [Copyright &y& Elsevier]

Abstract: Optimisation of biocatalytic systems for the electroreduction of molecular O2 in biofuel cell cathodes implies screening of the catalytic activity of enzyme/redoxpolymer assemblies. Os-complex modified electrodeposition polymers are suggested for linking bilirubin oxidase catalysed O2 reduction via an electron hopping sequence along the redox polymer to the electrode. They can be non-manually precipitated on electrode surfaces by electrochemically induced pH modulation. Cyclic voltammetry provides a good estimation of the electrocatalytic activity of a redox polymer/enzyme modified electrode surface. In addition, scanning electrochemical microscopy operating in redox competition mode (RC-SECM) supplies images of the spatial distribution of the biocatalytic activity. [Copyright &y& Elsevier]

• The importance of biosensor design and thorough characterization is presented. • In the presence of herbicides PSII still transfers electrons to the redox polymer. • Envisaged analytes may also affect other components of the electrode architecture. • Increased photocurrents are observed in the presence of a phenolic inhibitor. • Counterion effects have an important impact on redox polymer-based biosensors. Interfacing photosynthetic protein complexes with electrodes is frequently used for the identification of electron transfer mechanisms and the fabrication of biosensors. Binding of herbicide compounds to the terminal plastoquinone Q B at photosystem II (PSII) causes disruption of electron flow that is associated with a diminished performance of the associated biodevice. Thus, the principle of electron transport inhibition at PSII can be used for herbicide detection and has inspired the fabrication of several biosensors for this purpose. However, the biosensor performance may reveal a more complex behavior than generally expected. As we present here for a photobioelectrode constituted by PSII embedded in a redox polymer matrix, the effect caused by inhibitors does not only impact the electron transfer from PSII but also the properties of the polymer film used for immobilization and electrical wiring of the protein complexes. Incorporation of phenolic inhibitors into the polymer film surprisingly translates into enhanced photocurrents and, in particular cases, in a higher stability of the overall electrode architecture. The achieved results stress the importance to evaluate first the possible influence of analytes of interest on the biosensor architecture as a whole and provide important insights for consideration in future design of bioelectrochemical devices. [ABSTRACT FROM AUTHOR]

• Carbon nanoelectrodes etched in aqueous HF were modified with polymer/enzyme films. • Reliable fabrication of conical electrodes with a tip diameter of a few hundred nm. • Glucose oxidase was electrically wired by means of a redox polymer. • Polymer/enzyme modified electrodes could be used for intracellular measurements. • The biosensors showed a stable current output even after intracellular measurement. Carbon-based nanoelectrodes fabricated by means of pyrolysis of an alkane precursor gas purged through a glass capillary and subsequently etched with HF were modified with redox polymer/enzyme films for the detection of glucose at the single-cell level. Glucose oxidase (GOx) was immobilized and electrically wired by means of an Os-complex-modified redox polymer in a sequential dip coating process. For the synthesis of the redox polymer matrix, a poly(1-vinylimidazole- co -acrylamide)-based backbone was used that was first modified with the electron transfer mediator [Os(bpy) 2 Cl]+ (bpy = 2,2′-bipyridine) followed by the conversion of the amide groups within the acrylamide monomer into hydrazide groups in a polymer-analogue reaction. The hydrazide groups react readily with bifunctional epoxide-based crosslinkers ensuring high film stability. Insertion of the nanometre-sized polymer/enzyme modified electrodes into adherently growing single NG108-15 cells resulted in a positive current response correlating with the intracellular glucose concentration. Moreover, the nanosensors showed a stable current output without significant loss in performance after intracellular measurements. [ABSTRACT FROM AUTHOR]