Daren J. Caruana

Electrochemical immobilization of enzymes. Part V. Microelectrodes for the detection of glucose based on glucose oxidase immobilized in a poly(phenol) film

The adsorption of glucose oxidase at a platinum electrode followed by immobilization in an electrochemically polymerized phenol film was found to be a reproducible method for the fabrication of enzyme microelectrodes responsive to glucose. The observed responses were comparable to those predicted on the basis of monolayer coverage of the enzyme. The electrodes were found to be stable for more than 40 d on storage at 4 °C.

Enzyme-Amplified Amperometric Detection of Hybridization and of a Single Base Pair Mutation in an 18 Base Oligonucleotide on a 7 µm Diameter Microelectrode

A single base pair mismatch in an 18 base oligonucleotide was detected with a 7 µm diameter carbon microelectrode. The hybridization was followed directly and in real time by steady state amperometry. The microelectrode was coated with a hybridization-sensing layer in a two-step electrophoretic process, which yielded microelectrodes with reproducible dimensions. In the first step, a thin film of an electron-conducting redox polymer was deposited electrophoretically at constant potential in a low ionic strength solution. In the second step a carbodiimide activated single stranded probe was reactively electrophoretically deposited and covalently attached to the redox polymer film. The labeling enzyme, thermostable soybean peroxidase (SBP), was covalently bound to the 5’-end of the target single stranded oligonucleotide. When the redox polymer and the enzyme were brought to close proximity by hybridization of the target and probe oligonucleotides, the film on the electrode switched from being a non-catalyst to a catalyst for H2O2 electroreduction at -0.06 V vs. Ag/AgC1. The current observed corresponded to that generated by approximately 40,000 surface bound and electrically connected SBP molecules.

Electrochemistry of undoped diamond nanoparticles: accessing surface redox states.

The electrochemical response of an electrode-immobilized layer of undoped, insulating diamond nanoparticles is reported, which we attribute to the oxidation and reduction of surface states. The potentials of these surface states are pH-dependent; moreover they are able to interact with solution redox species. The voltammetric response of redox couples Fe(CN)(6)(3-/4-) and IrCl(6)(3-/2-) are compared at bare boron-doped diamond electrodes and electrodes modified with a layer of nanodiamond (ND). In all cases the presence of ND modifies the CV response at slow scan rates if low concentrations of redox couple are used. Enhancements of oxidation currents are noted at potentials at which the ND surface states can also undergo oxidation, and enhancements of reduction currents are likewise observed where ND is also reducible. We attribute these observations to electron transfer occurring between the species generated at the underlying electrode during CV and the ND immobilized in the interfacial region, leading to regeneration of the starting species and hence enhancement in currents due to a feedback mechanism. The magnitude of current enhancement thus depends on the standard potential of the redox couple relative to those of the ND surface states.

Modelling the response of a tungsten oxide semiconductor as a gas sensor for the measurement of ozone

The behaviour of gas-sensitive resistors based on WO3 towards small concentrations of ozone in air can be understood with a simple model involving the reaction of ozone with surface oxygen vacancies. This model has been validated by comparison with experimental results for the effects of varying oxygen partial pressure on the ozone response. A complete description of the behaviour of devices constructed by printing WO3 as porous layers onto an impermeable substrate requires consideration of the effects of the microstructure of such a device upon its response. A very simple series-parallel equivalent circuit model captures the effects and allows a simple interpretation of the sensor behaviour, including the quadratic limiting steady state resistance response to ozone and the effects of variation of device thickness. An important fact that allows WO3 to be used at rather high temperatures as an effective ozone sensor is that ozone does not decompose at any discernible rate on the oxide surface. Saturation of the oxide surface at ambient temperature with water vapour inhibits the ozone response when the sensor is subsequently heated. The effect can be removed by heating at sufficiently high temperature. Water vapour also gives a high-temperature sensor response, but appears to act at sites different to those that mediate the response to ozone.

Electrochemical immobilization of enzymes. Part VI. Microelectrodes for the detection of L-lactate based on flavocytochrome b2 immobilized in a poly(phenol) film

The adsorption of flavocytochrome b2 at a 25 µm diameter platinum microelectrode, followed by immobilization in an electrochemically polymerized phenol film, was found to be a reproducible method for the fabrication of microelectrodes responsive to L-lactate. Fabrication of enzyme microelectrodes in this manner is simple and produces electrodes that respond rapidly to L-lactate. The immobilization in the poly(phenol) film has no effect on the specificity of the enzyme for L-lactate over β-chlorolactate and reduces the direct electrochemical interference from uric acid, acetomidophenol and ascorbate. The stability of the resulting enzyme electrode is discussed.

Electron transfer mediated by glucose oxidase at the liquid/liquid interface

In order to establish an experimental basis for exploring the reactivity of membrane-bound redox enzymes using electrochemistry at an organic/aqueous interface, the reactivity of glucose oxidase adsorbed at the dichloroethane/water interface has been studied. Turnover of glucose in the aqueous phase mediated by dimethyl ferricenium electrogenerated in the organic phase was measured by measuring the feedback current caused by recycling the mediator as the generator electrode approached close to the interface from the organic side. An unexpected self-exchange reaction of the ferrocene at the interface was suppressed by adsorption of a surfactant. The interfacial enzyme reaction could be distinguished from reaction within the bulk of the aqueous phase. Reaction within a protein-surfactant film formed at the interface is conjectured.

Selective protein and DNA adsorption on PLL-PEG films modulated by ionic strength.

We describe a soft thin film which selectively adsorbs DNA but averts the non-specific binding of proteins. Indium tin oxide (ITO) substrates were surface-modified with a poly(L-lysine)-g-poly(ethylene glycol) (PLL-PEG) film which carries an outer protein-repelling PEG layer and an underlying positively charged PLL layer that attracts DNA. Binding of DNA could be tuned by a factor of over 90 by varying the salt concentration. The dependence of DNA binding on ionic strength was described with a physicochemical model which led to the conclusion of an unexpectedly high enrichment of salt inside the PEG layer. In addition, the model led to an expanded definition of the Debye-Huckel type effective screening length parameter z. Our new findings on a film with dual passivation/attraction properties can find applications in biopolymer-specific coatings useful in bioseparation and biosensing. In addition, the physicochemical characterisation provides new insight into the interactions between biopolymers and polymer-coated interfaces.

Self propagating high temperature synthesis of magnesium zinc ferrites (MgxZn1-xFe2O3): thermal imaging and time resolved X-ray diffraction experiments

Spinel ferrites of the form MgxZn1-xFe2O4 ( x = 0. 0.25, 0.50, 0.75, 1.00) were prepared by self-propagating high-temperature synthesis (SHS) from reactions of iron(III), zinc and magnesium oxides, iron powder and sodium perchlorate. The driving force for the reactions is the oxidation of iron powder. Reactions were carried out in the presence of an external magnetic field of 0.2 or 1.1 T. Reaction velocity and temperatures were obtained by thermal imaging camera. The transformation of reactants to products was studied by time resolved X-ray diffraction using Rietveld refinement for determination of phase percentages. Reactions typically reached temperatures in excess of 1150 degreesC with a timescale of complete conversion of reactant to products of 20 s. All materials were characterised by X-ray powder diffraction (XRD), energy dispersive X-ray analysis (EDXA), scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FTIR), Mossbauer spectroscopy and vibrating sample magnetometry (VSM).

Combustion synthesis of alkaline-earth substituted lanthanum manganites; LaMnO3, La0.6Ca0.4MnO3 and La0.6Sr0.4MnO3

Alkaline-earth substituted lanthanum manganites La1 − xAxMnO3 (A = Ca, Sr; x = 0, 0.4) were synthesized in air by self-propagating high-temperature synthesis (SHS): a combustion process involving the reaction of lanthanum(III) oxide, calcium oxide (or carbonate), strontium peroxide, manganese metal powder and sodium perchlorate. A very sensitive thermal imaging method was used to study the combustion characteristics of the SHS-process. This method is based on the continuous measurement of the combustion process using an IR-camera and software developed by MIKRON Instrument Co., Inc. (M9100 Pyrovision Series – Imaging Pyrometer). X-Ray powder diffraction, scanning electron microscopy (SEM)/energy dispersive analysis by X-rays (EDAX), infrared spectroscopy and vibrating sample magnetometry were carried out on all samples. X-Ray diffraction data showed that single-phase orthorhombic LaMnO3 and monoclinic La0.6Ca0.4MnO3 and La0.6Sr0.4MnO3 were formed. Magnetic properties were found to transform from antiferromagnetic for LaMnO3 to weak ferromagnetic for La0.6Ca0.4MnO3 to strong ferromagnetic for La0.6Sr0.4MnO3. The experimental results show the SHS route is useful for the synthesis of multicomponent Mn-based oxides. The thermal imaging work enables accurate monitoring of the SHS combustion process and enabled velocity of propagation, maximum temperature, cooling rates, synthesis wave width and synthesis wave path to be determined with high precision.

Low-temperature treatment of semiconducting interlayers for high-efficiency light-emitting diodes based on a green-emitting polyfluorene derivative

We investigate the scope for low-temperature treatment of exciton/electron blocking interlayers in light-emitting diodes based on poly(9,9′-dioctylfiuorene-alt-benzothiadiazole) (F8BT). We focus on poly(9,9′-dioctylfluorene-alt-N-(4-butylphenyl)-diphenylamine) (TFB) interlayers processed at temperatures up to 50 °C, i.e., far below the glass transition temperature of TFB (∼156 °C). Continuous-wave and time-resolved photoluminescence studies confirm the formation of both excitons and exciplex species, as a result of the F8BT/TFB intermixing. Interestingly, however, we can still increase the electroluminescence external quantum efficiency from 0.05% to 0.5% and 1% for progressively thicker TFB films. We propose that a degree of intermixing may become acceptable as a trade-off to achieve low-temperature processability.