Edmond Magner

Proteins in Mesoporous Silicates

Mesoporous silicates (MPS) have an ordered pore structure with dimensions comparable to many biological molecules. They have been extensively explored as supports for proteins and enzymes in biocatalytic applications. Since their initial discovery, novel syntheses methods have led to precise control over pore size and structure, particle size, chemical composition, and stability, thus allowing the adsorption of a wide variety of biological macromolecules, such as heme proteins, lipases, antibody fragments, and proteases, into their structures. This Review discusses the application of ordered, large-pore, functionalized mesoporous silicates to immobilize proteins for biocatalysis.

Self-powered wireless carbohydrate/oxygen sensitive biodevice based on radio signal transmission

Here for the first time, we detail self-contained (wireless and self-powered) biodevices with wireless signal transmission. Specifically, we demonstrate the operation of self-sustained carbohydrate and oxygen sensitive biodevices, consisting of a wireless electronic unit, radio transmitter and separate sensing bioelectrodes, supplied with electrical energy from a combined multi-enzyme fuel cell generating sufficient current at required voltage to power the electronics. A carbohydrate/oxygen enzymatic fuel cell was assembled by comparing the performance of a range of different bioelectrodes followed by selection of the most suitable, stable combination. Carbohydrates (viz. lactose for the demonstration) and oxygen were also chosen as bioanalytes, being important biomarkers, to demonstrate the operation of the self-contained biosensing device, employing enzyme-modified bioelectrodes to enable the actual sensing. A wireless electronic unit, consisting of a micropotentiostat, an energy harvesting module (voltage amplifier together with a capacitor), and a radio microchip, were designed to enable the biofuel cell to be used as a power supply for managing the sensing devices and for wireless data transmission. The electronic system used required current and voltages greater than 44 µA and 0.57 V, respectively to operate; which the biofuel cell was capable of providing, when placed in a carbohydrate and oxygen containing buffer. In addition, a USB based receiver and computer software were employed for proof-of concept tests of the developed biodevices. Operation of bench-top prototypes was demonstrated in buffers containing different concentrations of the analytes, showcasing that the variation in response of both carbohydrate and oxygen biosensors could be monitored wirelessly in real-time as analyte concentrations in buffers were changed, using only an enzymatic fuel cell as a power supply.

Adsorption and Activity of Proteins onto Mesoporous Silica

The adsorption and activity of cytochrome c onto two different MCM-41 materials, MCM-41/28 and MCM-41/45 with average pore diameters of 28 and 45 Å respectively, is presented. Nitrogen gas adsorption/desorption isotherms before and after protein adsorption, and peroxidative activity profiles of the adsorbed protein demonstrate that the protein is adsorbed into the mesoporosity and remains active. The adsorption of a range of different proteins onto both MCM-41/28 and 45 shows how protein properties affect adsorption.

Trends in electrochemical biosensors

The development of electrochemically based biosensors is discussed in the context of what has been learned from the successful development of glucose biosensors. Some future trends are discussed.

Characterization of Nanoporous Gold Electrodes for Bioelectrochemical Applications

The high surface areas of nanostructured electrodes can provide for significantly enhanced surface loadings of electroactive materials. The fabrication and characterization of nanoporous gold (np-Au) substrates as electrodes for bioelectrochemical applications is described. Robust np-Au electrodes were prepared by sputtering a gold-silver alloy onto a glass support and subsequent dealloying of the silver component. Alloy layers were prepared with either a uniform or nonuniform distribution of silver and, post dealloying, showed clear differences in morphology on characterization with scanning electron microscopy. Redox reactions under kinetic control, in particular measurement of the charge required to strip a gold oxide layer, provided the most accurate measurements of the total electrochemically addressable electrode surface area, A(real). Values of A(real) up to 28 times that of the geometric electrode surface area, A(geo), were obtained. For diffusion-controlled reactions, overlapping diffusion zones between adjacent nanopores established limiting semi-infinite linear diffusion fields where the maximum current density was dependent on A(geo). The importance of measuring the surface area available for the immobilization was determined using the redox protein, cyt c. The area accessible to modification by a biological macromolecule, A(macro), such as cyt c was reduced by up to 40% compared to A(real), demonstrating that the confines of some nanopores were inaccessible to large macromolecules due to steric hindrances. Preliminary studies on the preparation of np-Au electrodes modified with osmium redox polymer hydrogels and Myrothecium verrucaria bilirubin oxidase (MvBOD) as a biocathode were performed; current densities of 500 μA cm(-2) were obtained in unstirred solutions.

Characterization of β-Lactoglobulin Fibrillar Assembly Using Atomic Force Microscopy, Polyacrylamide Gel Electrophoresis, and in Situ Fourier Transform Infrared Spectroscopy

The aggregation process of beta-lactoglobulin (beta-lg) from 0 min to 20 h was studied using atomic force microscopy (AFM), scanning transmission electron microscopy (STEM), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and in situ attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR). Fibril assembly was monitored in real time using AFM up to 20 h. From 0 to 85 min, beta-lg monomers deformed and expanded with some aggregation. After 85 min, fibrillar structures were formed, exceeding 10 mum in length. Fibrillar structures were confirmed by STEM. Secondary structural changes occurring during fibril formation were monitored by ATR-FTIR at 80 degrees C and indicated a decrease in alpha-helix content and an increase in beta-sheet content. SDS-PAGE indicated that fibrils were composed of polypeptides and not intact monomers. In this study, beta-lg and whey protein isolate (WPI)-derived fibrils, including some double helices, in water were observed by AFM under ambient conditions and in their native aqueous environment.

Conductive copolymer-modified carbon fibre microelectrodes: electrode characterisation and electrochemical detection of p-aminophenol

Abstract Thin film electro-coated poly[N-vinylcarbazole-co-vinylbenzene sulfonic acid] (p[NVCzVBSA]), poly[carbazole-co-methylthiophene] (p[CzMeTh]) and polycarbazole (p[Cz]) carbon fibre microelectrodes (CFMEs) were characterised by scanning electron microscopy (SEM) and FTIR-ATR spectroscopy. These modified carbon fibre electrodes were found to be effective systems for the determination of para-aminophenol (p-AP). Thin film coated p[NVCzVBSA] was the most suitable modified electrode for the detection of p-AP.

Adsorption and activity of cytochrome c on mesoporous silicates

Cytochrome c (horse heart) has been adsorbed onto a range of mesoporous silicate materials with the extent of adsorption dependent on the silicate pore size; adsorption and activity profiles of the adsorbed protein are reported.

Direct electron transfer of trametes hirsuta laccase adsorbed at unmodified nanoporous gold electrodes

The enzyme Trametes hirsuta laccase undergoes direct electron transfer at unmodified nanoporous gold electrodes, displaying a current density of 28μA/cm(2). The response indicates that ThLc was immobilised at the surface of the nanopores in a manner which promoted direct electron transfer, in contrast to the absence of a response at unmodified polycrystalline gold electrodes. The bioelectrocatalytic activity of ThLc modified nanoporous gold electrodes was strongly dependent on the presence of halide ions. Fluoride completely inhibited the enzymatic response, whereas in the presence of 150mM Cl(-), the current was reduced to 50% of the response in the absence of Cl(-). The current increased by 40% when the temperature was increased from 20°C to 37°C. The response is limited by enzymatic and/or enzyme electrode kinetics and is 30% of that observed for ThLc co-immobilised with an osmium redox polymer.

A facile aqueous sol–gel method for high surface area nanocrystalline CeO2

Thermally stable high surface area nanocrystalline CeO2 was synthesized via an aqueous sol–gel method using cerium hydroxide and urea as precursors. The CeO2 sol is yellow colored and absorbs light over the range 200–500 nm. Nanocrystalline CeO2 particles derived from the sol are spherical in shape and range from 9 to 13 nm in diameter. The CeO2 powder has a high surface area of up to 85 m2 g−1 and a band gap of 3.1 eV. The method of preparation is facile, utilises low cost reagents and can be performed on a large scale.