The development and characterisation of some novel amperometric sensors

Chapter 1 of this thesis serves as a general introduction to electrode processes and to some of the electrochemical techniques used for the characterisation and development of amperometric sensors. A brief review of recent applications of modified electrodes in the field of electroanalysis is also presented. The remainder of the thesis is divided into three sections. The first of these comprises chapters 2 and 3. Chapter 2 describes the synthesis and characterisation of a series of polymers based on ruthenium-bound 4-vinylpyridine/styrene copolymers. An investigation into factors that affect charge transport rates and processes through thin films of these polymers on glassy carbon electrodes, using cyclic voltammetry, chronoamperometry and sampled current voltammetry, is also presented. Chapter 3 describes the optimisation of the performance of these polymer modified electrodes as sensors for the nitrite ion, using the rotating disk electrode and flow-injection amperometric detection techniques. The second section of this thesis comprises the development of ruthenium dioxide-based modified electrodes for the electrocatalytic detection of alcohols and selected saccharide antibiotics. The ruthenium dioxide modifier is also confined in a graphite-epoxy matrix to yield a stable, polishable sensor for the saccharide antibiotics. Finally, chapter 5 describes the development of, and investigation into, two sensors for the detection of glucose. The first of these is based on the electrocatalytic oxidation of glucose at a ruthenium dioxide-modified graphite-epoxy electrode surface, while the second involves a novel one-step fabrication of a glucose sensor based on the entrapment of the enzyme glucose oxidase within a poly(ester sulfonic acid) coating

A Comparative Study between Hydrogen Peroxide Amperometric Biosensors Based on Different Peroxidases Wired by Os-Polymer: Applications in Water, Milk and Human Urine

In the last few years, hydrogen peroxide [...

A Comparative Study between Hydrogen Peroxide Amperometric Biosensors Based on Different Peroxidases Wired by Os-Polymer: Applications in Water, Milk and Human Urine

In the last few years, hydrogen peroxide [...

Sensors based on polymer modified electrodes

This paper will review the recent results that we have obtained using novel ruthenium-containing polymers, and on the further studies on the incorporation of proteins into polymeric matrices

Nitrogenase bioelectrocatalysis: heterogeneous ammonia and hydrogen production by MoFe protein

notrogenase is the only enzyme known to catalyze the reduction of N2 to 2NH3. In vivo, the MoFe protein component of nitrogenase is exclusively reduced by the ATP-hydrolyzing Fe protein in a series of transient association/dissociation steps that are linked to the hyderolysis of two ATP for each electron transeferred. We report MoFe protein immobilized at an electrode surface, where cobaltocene (as an electron mediator that can be observed in real time at a carbon electrode) is used to reduce the MoFe protein (independent of the Fe protein and of ATP hydrolysis) and support the bioelectrocatalytic reduction of protons to dihydrogen, azide to ammonia, and nitrit to ammonia. Bulk bioelectrosynthetic N3 or NO2 reduction (50 mM) for 30 minutes yielded 70 +- 9 nmol NH3 and 234 +- 62 nmol NH3, with NO2 reduction operating at high faradaic efficiency

Designing stable redox-active surfaces: chemical attachment of an osmium complex to glassy carbon electrodes prefunctionalized by electrochemical reduction of an in situ-generated aryldiazonium cation.

International audienceThe production of stable redox-active layers on electrode surfaces can lead to improvements in electronic device design. Enhanced stability can be achieved by pretreatment of electrode surfaces to provide surface chemical functional groups for covalent tethering of redox complexes. Herein, we describe pretreatment of glassy carbon electrodes to provide surface carboxylic acid groups by electro-reduction of an in situ-generated aryl diazonium salt from 3-(4-aminophenyl)propionic acid. This surface layer is characterized by attenuated total reflection infrared spectroscopy, atomic force microscopy, and electrochemical blocking studies. The surface carboxylic acid generated is then used to tether an osmium complex, [Os(2,2'-bipyridyl)2(4-aminomethylpyridine)Cl]PF6, to provide a covalently bound redox-active monolayer, E(0) ' of 0.29 V (vs Ag/AgCl in phosphate buffer, pH 7.4), on the pretreated glassy carbon electrode. The layer proves stable to pH, temperature, and storage conditions, retaining electroactivity for at least 6 months

Increasing Redox Potential, Redox Mediator Activity, and Stability in a Fungal Laccase by Computer-Guided Mutagenesis and Directed Evolution

Fungal high-redox-potential laccases (HRPLs) are multicopper oxidases with a relaxed substrate specificity that is highly dependent on their binding affinity and redox potential of the T1Cu site (ET1). In this study, we combined computational design with directed evolution to tailor an HRPL variant with increased ET1 and activity toward high-redox-potential mediators as well as enhanced stability. Laccase mutant libraries were screened in vitro using synthetic high-redox-potential mediators with different oxidation routes and chemical natures, while computer-aided evolution experiments were run in parallel to guide benchtop mutagenesis, without compromising protein stability. Through this strategy, the ET1 of the evolved HRPL increased from 740 to 790 mV, with a concomitant improvement in thermal and acidic pH stability. The kinetic constants for high-redox-potential mediators were markedly improved and were then successfully tested within laccase mediator systems (LMSs). Two hydrophobic substitutions surrounding the T1Cu site appeared to underlie these effects, and they were rationalized at the atomic level. Together, this study represents a proof-of-concept of the joint elevation of the ET1, redox mediator activity, and stability in an HRPL, making this versatile biocatalyst a promising candidate for future LMS applications and for the development of bioelectrochemical devices.This work was supported by the European Union project Bioenergy-FP7-PEOPLE-2013-ITN-607793, the COST Action [CM1303 Systems Biocatalysis],the Swedish Energy Agency (44707-1), the Knowledge Foundation (20170168), and the Spanish Government [BIO2016-79106-R-Lignolution and CTQ2016-79138-R].We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI)