No description available.

No description available.

An alternative use for an amperometric enzyme electrode is as an anodic half-cell of a fuel cell. A biofuel cell based upon the oxidation of methanol by the quinoprotein alcohol dehydrogenase was developed.

abstract / Biological Sciences / Doctoral / Doctor of Philosophy

No description available.

Today’s most pressing problems would greatly benefit from an integrated production method for food, water, and energy. Biological fuel cells can offer such a production method, but current designs cannot be scaled to meet global demand. The ability of five different fungal strains to secrete laccase was evaluated under optimized culture conditions using three inducers. A specialized electrode was developed to increase the loading of laccase on the cathode. Trametes versicolor was then immobilized at the modified cathode and shown to secrete electrochemically active laccase. This hybrid design combines the power density of an enzymatic catalyst with the robustness of a microbial catalyst by facilitating biological renewal of the enzymatic catalyst laccase.

Ammonia is an important compound to many industries around the world. Most of the fertilizers used by crop growers have ammonia as an essential ingredient. It can also be useful as a fuel source, offering greater energy density per unit than hydrogen and greater safety. Currently, the predominant method for producing ammonia on an industrial scale is by the Haber-Bosch process. This process uses steam evolution of methane to provide H2 gas, which is then combined with N2 gas over an iron catalyst to form NH3. This process requires large amounts of energy as well as high temperatures and pressures. Here, an alternative method for ammonia production is explored. With Anabaena Variabilis, a photosynthetic cyanobacteria, on a carbon electrode, ammonia can be generated at ambient temperatures and pressures at little energy cost, a few tenths of a volt. A bioelectrocatalytic device has been constructed by immobilizing whole cell a. variabilis in a Nafion film modified with a trimethyl octadecyl ammonium bromide (TMODA) salt at an electrode surface [3]. The polymer modified electrode provides the driving force and reductive microenvironment to facilitate production of NH3 by nitrogenase and nitrate/nitrite reductase enzymes present in a. variabilis. Ammonia production by cyanobacteria were increased from basal levels of 2.8 ± 0.4 µM produced over a two week period, to 22 ± 8 µM produced in 20 minutes under mild voltage perturbation, roughly 104% increase in rate. Control of ammonia producing structures (nitrogenase in heterocystic cells or nitrate/nitrite reductase in vegetative cells) can be accomplished by growing the algae with and without fixed sources of nitrogen in the growth media. With the addition of various nitrogen-containing gases to the electrolyte solution during cyclic voltammetry, there is evidence that biofilms containing a mixture of cell types increases ammonia production above controls when the nitrogen is present as NO2-, NO, or N2O. Chronoamperometric perturbation studies show increased ammonia production at near +600 mV and -300 mV vs SCE. In cyclic voltammetric studies, nitrate/nitrite reductase in vegetative-only biofilms responds favorably to positive voltage ranges, while isolated heterocyst biofilms containing nitrogenase can be effectively targeted with the application of a negative voltage profile. References: [1] Johna Leddy and Timothy M. Pashkewitz, Ammonia Production Using Bioelectrocatalytic Devices, US Patent Application 20140011252 [2] Timothy M. Paschkewitz, Ammonia Production at Ambient Temperature and Pressure: An Electrochemical and Biological Approach, Ph.D., University of Iowa, 2012.

This dissertation explores the electrochemical behavior of enzymes and their cofactors at carbon nanotube (CNT) and nitrogen-doped carbon nanotube (N-CNT) electrodes. Two common types of oxidoreductases are considered: flavin adenine dinucleotide (FAD)-dependent oxidases and nicotinamide adenine dinucleotide-dependent (NAD⁺)-dehydrogenases. Chapter 1 presents the oxygen reduction reaction (ORR) at N-CNT electrodes as a way to electrochemically measure enzymatic turnover at the electrode surface. The unique peroxide pathway at N-CNT electrodes, which catalytically disproportionates hydrogen peroxide (H₂O₂) back into oxygen, provides an increased ORR current directly proportional to the rate of enzymatic turnover for H₂O₂ producing enzymes, even in an oxygen saturated solution. Biosensing of L-lactate using the increased ORR current is demonstrated using L-lactate oxidase. Chapter 2 explores the surface bound electrochemical signal of FAD when FAD-dependent enzyme or free FAD is allowed to spontaneously adsorb onto the CNT/N-CNT surface. Specifically, the origin of the enzymatically generated FAD signal and the rate constant of the electron transfer are elucidated. Chapter 3 continues the discussion of the cofactor FAD by demonstrating its use as an informative surface specific redox probe for graphitic carbon surfaces. Primarily, FAD can be used to determine the electroactive surface area and the relative hydrophobicity/hydrophilicity of graphitic surfaces. Chapter 4 changes gears to NAD⁺-dependent dehydrogenases by investigating the electrocatalytic oxidation of NADH at N-CNTs in comparison with conventional carbon electrodes or nondoped CNTs. Biosensing of glucose through the oxidation of NADH is demonstrated using glucose dehydrogenase adsorbed onto the N-CNT surface. Chapter 5 continues the discussion of NAD⁺-dependent dehydrogenases by addressing the reaction kinetics of NADH oxidation at N-CNTs as a tool to measure the enzymatic reduction of NAD⁺.

The aim of this thesis is to enhance the extracellular electron transfer rates (exoelectrogenesis) in cyanobacteria, to be utilised for photo-bioelectricity generation in biophotovoltaics (electrochemical cell). An initial cross comparison of the cyanobacterium Synechococcus elongatus PCC7942 against other exoelectrogenic cultures showed a hindered exoelectrogenic capacity. Nonetheless, in mediatorless biophotovoltaics, it outperformed the microalgae Chlorella vulgaris. Furthermore, the performance of S. elongatus PCC7942 was improved by constructing a more efficient design (lower internal resistance), which was fabricated with carbon fibres and nitrocellulose membrane, both inexpensive materials. To strategically obtain higher exoelectrogenic rates, S. elongatus PCC7942 was conditioned by iron limitation and CO2 enrichment. Both strategies are novel in improving cyanobacteria exoelectrogenesis. Iron limitation induced unprecedented rates of extracellular ferricyanide reduction (24-fold), with the reaction occurring favourably around neutral pH, different to the cultural alkaline pH. Iron limited cultures grown in 5% and 20% CO2 showed increased exoelectrogenic rates in an earlier stage of growth in comparison to air grown cultures. Conveniently, the cultural pH under enriched CO2 was around neutral pH. Enhanced photo-bioelectricity generation in ferricyanide mediated biophotovoltaics was demonstrated. Power generation was six times higher with iron limited cultures at neutral pH than with iron sufficient cultures at alkaline pH. The enhanced performance was also observed in mediatorless biophotovoltaics, especially in the dark phase. Exoelectrogenesis was mainly driven by photosynthetic activity. However, rates in the dark were also improved and in the long term it appeared that the exoelectrogenic activity under illumination tended to that seen in the dark. Proteins participating in iron uptake by an alleged reductive mechanism were overexpressed (2-fold). However, oxidoreductases in the outer membrane remain to be identified. Furthermore, electroactive regions in biofilms of S. elongatus PCC7942 were established using cyclic voltammetry. Double step potential chronoamperometry was also successfully tested in the biofilms. Thus, the electrochemical characterisation of S. elongatus PCC7942 was demonstrated, implying that the strategies presented in this thesis could be used to screen for cyanobacteria and/or electrode materials to further develop systems for photo-bioelectricity generation.

Esta tesis doctoral se centra en el desarrollo de diferentes métodos electroquímicos y químicos para la funcionalización de materiales de carbono nanoestructurados, obteniendo materiales electródicos funcionales para aplicaciones bioelectroquímicas. En este sentido, las propiedades en superficie influyen directamente en la interacción entre el elemento bioreceptor o biocatalizador con el electrodo. Por tal motivo, se puede mejorar la cinética de transferencia de electrones, la inmovilización, orientación y distribución del bioelemento en el electrodo, mejorando el rendimiento del dispositivo bioelectroquímico, a partir de la química superficial del material empleado. El uso de diferentes funcionalidades de nitrógeno y fósforo generados electroquímicamente sobre los materiales de carbono nanoestructurados proporciona una plataforma para la síntesis controlada, para mejorar la actividad catalítica de biocatalizadores. Además, el proceso electroquímico de funcionalización ha demostrado ser una ruta interesante para preparar bioelectrodos en un solo paso, con bajo consumo de elementos enzimáticos y un rendimiento sobresaliente a los métodos convencionales actuales. Por otro lado, el uso de materiales de carbono dopados en N cuaternarios como elemento transductor en la síntesis de un biosensor enzimático de glucosa libre de metales proporciona, en condiciones aeróbicas, un biosensor de alta sensibilidad para la detección de glucosa en orina y bebidas azucaradas comerciales, con bajo efecto de los interferentes y alta estabilidad. Finalmente, la modificación de los nanotubos de carbono con nanopartículas de oro proporciona un material de electrodo transductor escalable y estable en el que tiene lugar la inmovilización de anticuerpos a través de interacciones Au-8. La formación de antígeno-anticuerpo complejo provoca efectos estéricos que producen un impedimento para la transferencia de electrones proveniente de una sonda redox activa hacia la superficie del electrodo, lo que facilita la detección del analito.