Electroenzymatic Processes Research Articles

Electro-enzymatic processes in mitochondria

Four enzymatic complexes form the electron transport chain of the mitochondrion. Complexes I and II are current generator half-elements, that create electron flows with the participation of the enzymes NADH dehydrogenase and Succinate dehydrogenase, oxidizing NADH·H into NAD+, and succinate into fumarate, respectively. Electrons are transferred throw the iron-sulfur clusters system, participate reduction enzymatic reaction of CoQ10 into CoQ10Н2. The proton pumping into the mitochondrial intermembrane space, as well as the regeneration of CoQ10 occur when electrons transferred by ubiquinone-cytochrome-c-oxidoreductase enzyme to the Riske iron-sulfur protein (Fe2S2). The Riske protein regulates electron flows: it transfers one electron to the bL,bН (III) complex, and the other to cytochrome c. Cytochrome c directs electrons to the ΙV complex electrolysis system. The binuclear copper center [CuA-CuB] of the ΙV complex is the anode, where oxygen is formed. Electrons along the galvanic pair chain from paired copper Cu2++ e- ↔ Cu+ and iron Fe2+- е- ↔Fe3+ are transferred to the cathode, copper ion (Сu+), where the electrochemical reaction occurs O2 + 4e- + 4H+ → 2H2O, with the pumping of 4 protons into the mitochondrial intermembrane space.

Quantitative and Non‐Quantitative Assessments of Enzymatic Electrosynthesis: A Case Study of Parameter Requirements

AbstractThe integration of enzymatic and electrochemical reactions offers a unique opportunity to optimize production processes. Recently, an increasing number of laboratory‐scale enzymatic electrosyntheses have shown impressive performance indicators, leading to scientific interest in technical implementation. However, important process parameters are missing in most of the relevant literature. On one hand, this is due to the large variety of relevant performance indicators. On the other hand, enzyme technologists and electrochemists use different parameters to describe a process. In this article, we review the most important performance indicators in electroenzymatic processes and suggest that in order to allow quantitative comparison, these indicators should be reported in all respective publications. In addition to quantitative parameters, non‐quantitative assessments often need to be included in a final evaluation. Examples of such parameters are sustainability, contribution to the UN Sustainable Development Goals or interactions with the overall process. We demonstrate the evaluation of processes using hydrogen peroxide‐dependent peroxygenases. The strength of the proposed evaluation system lies in its ability to identify weaknesses in a process at an early stage of development. Finally, it can be concluded that all evaluated enzymatic electrosynthesis do not yet meet typical industrial requirements for an enzyme‐based process.

Selectivity and Sustainability of Electroenzymatic Process for Glucose Conversion to Gluconic Acid

Electroenzymatic processes are interesting solutions for the development of new processes based on renewable feedstocks, renewable energies, and green catalysts. High-selectivity and sustainability of these processes are usually assumed. In this contribution, these two aspects were studied in more detail. In a membrane-less electroenzymatic reactor, 97% product selectivity at 80% glucose conversion to gluconic acid was determined. With the help of nuclear magnetic resonance spectroscopy, two main side products were identified. The yields of D-arabinose and formic acid can be controlled by the flow rate and the electroenzymatic reactor mode of operation (fuel cell or ion-pumping). The possible pathways for the side product formation have been discussed. The electroenzymatic cathode was found to be responsible for a decrease in selectivity. The choice of the enzymatic catalyst on the cathode side led to 100% selectivity of gluconic acid at somewhat reduced conversion. Furthermore, sustainability of the electroenzymatic process is estimated based on several sustainability indicators. Although some indicators (like Space Time Yield) are favorable for electroenzymatic process, the E-factor of electroenzymatic process has to improve significantly in order to compete with the fermentation process. This can be achieved by an increase of a cycle time and/or enzyme utilization which is currently low.

Towards electroenzymatic processes involving old yellow enzymes and mediated cofactor regeneration.

Old yellow enzymes are able to catalyze asymmetric C=C reductions. A mediated electroenzymatic process to regenerate the NADPH in combination with an old yellow enzyme was investigated. Due to the fact that the overall process was affected by a broad set of parameters, a design of experiments (DoE) approach was chosen to identify suitable process conditions. Process conditions with high productivities of up to 2.27 mM/h in combination with approximately 90% electron transfer efficiency were identified.

Enantioselective enzymatic synthesis of the α-hydroxy ketone (R)-acetoin from meso-2,3-butanediol

Acetoin (3-hydroxy-2-butanone) is an important flavour compound and is applied in cosmetics, pharmacy and chemical synthesis. In contrast to chemical syntheses or fermentations an enzymatic route facilitates enantioselective acetoin production. The discovery of a (S)-selective alcohol dehydrogenase enables a novel production process of (R)-acetoin from meso-2,3-butanediol. It was shown that the regeneration of oxidised nicotinamide adenine dinucleotide is a key point in preparative application of dehydrogenases for the oxidative route. An electrochemical regeneration system was successful combined with the ADH catalysed reaction. Up to 48 mM (R)-acetoin was produced in the reaction system while productivities up to 2 mM h−1 were reached. The possibility to apply an electrochemical system in a semi-preparative synthesis will stimulate further research of electroenzymatic processes with oxidoreductases.

Coupling of electrochemical and optical measurements in a microtiter plate for the fast development of electro enzymatic processes with P450s

Electro enzymatic processes offer novel opportunities in catalysis by combining advantages of enzyme catalysis and electrochemistry. An efficient electrochemical cofactor substitution system can help to overcome economical hurdles for the technical use of cofactor dependent enzymes. The in vitro biocatalysis with P450 BM-3 was investigated aiming for the substitution of the expensive natural cofactor NADPH by electrochemistry as “electron source”. An electrochemical 24-well microtiter plate (eMTP) was developed, which can be employed in a standard microtiter plate reader and enables parallelized electrochemical experiments in combination with simultaneous optical measurements. The eMTP was applied to screen a P450 monooxygenase BM-3 mutein library and determine the behavior of P450 BM-3 muteins in an electrochemically driven surrogate assay with the mediator cobalt sepulchrate. Besides determining reaction rates also the influence of single reaction parameters e.g. applied potential, enzyme and mediator concentration were measured. Additionally the developed eMTP based screening system allows a fast development of an electro enzymatic process.

Electro-enzymatic degradation of chlorpyrifos by immobilized hemoglobin

Electro-enzymatic processes, which are enzyme catalysis combined with electrochemical reactions, have been used in the degradation of many environment pollutants. For some pollutants, the catalytic mechanisms of the electrochemical-enzyme reaction are still poorly understood. In this paper, the degradation of chlorpyrifos by a combination of immobilized hemoglobin and in situ generated hydrogen peroxide is reported for the first time. Hemoglobin was immobilized on graphite felts to catalyze the removal of chlorpyrifos in an electrochemical-enzyme system. Under the optimal conditions, more than 98% of the chlorpyrifos was degraded. Furthermore, the degradation products of chlorpyrifos were also studied and identified using liquid chromatography-mass spectrometry analysis. The results suggest a possible degradation mechanism for chlorpyrifos with low power and high efficiency, reveal the feasibility of hemoglobin as a substitute for some expensive natural enzymes, and demonstrate the application of an electro-enzymatic process in the treatment of organophosphorus compounds in wastewater.

Stable Electroenzymatic Processes by Catalyst Separation

Division of labour: The rapid enzyme inactivation in the electroenzymatic synthesis of chiral alcohols has been the main obstacle for synthetic applications during the last two decades. The reasons for this inactivation have now been elucidated. The development of a water-soluble polymeric mediator and the spatial separation of enzyme and mediator led to the first stable process and significantly improved catalyst utilisations (see picture).

Productivity of Selective Electroenzymatic Reduction and Oxidation Reactions: Theoretical and Practical Considerations

AbstractThe volumetric productivities, final product concentration and total process times, of electroenzymatic processes comprising reductive electrochemical cofactor regeneration coupled to dehydrogenase or oxygenase catalyzed redox reactions determine process performance. Exemplified for the production of fine chemicals, operational windows were defined to consider these parameters. This theoretical approach allows the identification of limiting process parameters and promising process developments. Several biocatalytic processes for syntheses of specialty chemicals are already in a performance range of technical relevance. Electroenzymology thus evolved to a strong additional technology in the toolbox of biocatalysis and organic chemistry.

Chronopotentiometric study of electroenzymatic processes with homogeneous mediation

A chronopotentiometric study for the system glucose-glucose oxidase in the presence of diaminodurene as an electron mediator is presented. The changes of the anodic overpotential of the mediator with time during constant current oxidation or used to discuss the kinetics in an electroenzymatic system with homogenous mediation. The shape of the chronopotentiograms is strongly dependent on the current range and for currents close to the diffusion limiting current of the mediator ( I → I d), the potential approaches a time-independent value. As the current increases, the shape of the chronopotentiograms changes to a sigmoid, with a transition time (τ) larger than the corresponding diffusion transition time ( τ d) of the free mediator. The limiting current of the mediator-enzyme-substrate system ( I lim) is considerably larger than I d. The ratio Iτ 1 2 Iτ d 1 2 decreases with increasing currents, approaching a constant value as I → I lim. The observed features are explained in terms of a theoretical model, which also enables an estimate of the rate constant for the catalytic process.

Electroenzymatic Processes: A Clean Technology Alternative for Highly Selective Synthesis?

In many fields such as fine chemistry and pharmacy, biotechnology is increasingly being used to produce natural-like molecules of great interest. The main method remains fermentation which results in complex mixtures in which there is only a very small amount of the required compound. Numerous downstream separation steps are necessary, which are expensive and generate large amounts of polluting wastes such as solvents, resins or salts. Using free enzymes extracted from microorganisms would avoid most of these separations and wastes. Oxidoreductases catalyse a lot of electron transfer reactions involved in many economically interesting syntheses, but electron extraction or feed is still a critical stage in developing free enzyme processes with them. The use of an electrochemical reactor to ensure electrical extraction or feed would provide a good step towards cleaner processes because the highest selectivity would be ensured without any by-products being generated. The first purpose of this paper is to show the broad application field of oxidoreductases in which electrochemistry could be introduced with some benefit. A concise analysis of the first electroenzymatic reactor design shows that in order to develop such economically efficient reactors, the enzymes must be confined in the strict vicinity of the electrode surface. Consequently, the experimental part is devoted to the development of an electrode/enzyme interface specificially adapted to synthesis processes. Electropolymerisation of pyrrole on a platinum electrode was used to entrap successively glucose oxidase for gluconic acid production, and hydrogenase for NADH regeneration. Emphasis was placed on increasing the mass transfer rates in the film and the amount of enzyme retained. © 1997 SCI.

Electroenzymatic Processes: A Clean Technology Alternative for Highly Selective Synthesis?

Electroenzymatic Processes: A Clean Technology Alternative for Highly Selective Synthesis?

A kinetic model for the electroenzymatic processes involved in polyphenol-oxidase-based amperometric catechol sensors

A kinetic model is presented for the electroenzymatic processes occurring at a polyphenol oxidase rotating-disk-bioelectrode for low concentrations of catechol. The work presented here is based on the same formalism as that previously used by Bartlett and Whitaker (J. Electroanal. Chem., 224 (1987) 27) for a glucose sensor. Our model allows one to analyze the effect of enzyme loading, film thickness, permeability and electrode rotation rate. It also unambiguously demonstrates for the first time that the high sensitivity and low detection limit observed for such an enzymatic electrode is the consequence of the electrochemical recycling of substrate which occurs upon transduction.