Rapid fingerprinting of bacterial species using nanocavities created on screen-printed electrodes modified by ß-cyclodextrin.

Rapid and precise identification of infectious microorganisms is important across a range of applications where microbial contamination can cause serious issues ranging from microbial resistance to corrosion. In this paper a screen-printed, polymeric ß-cyclodextrin (ß-CD) modified electrode, affording nanocavities for inclusion of the analytes, is shown as a disposable sensor capable of identifying bacteria by their metabolites. Three bacterial species were tested: two from the Pseudomonas genus, Pseudomonas fluorescens (P. fluorescens) and Pseudomonas aeruginosa (P. aeruginosa), and Serratia marcescens (S. marcescens), a member of the family, Enterobacteriaceae. On biofilm formation each species gave distinct, reproducible, redox fingerprints with a detection limit of 4 × 10-8 M. Square wave adsorptive stripping voltammetry (SWAdSV) was used for detection. Scanning electron microscopy (SEM) and cyclic voltammetry (CV) techniques were used to characterize the morphology and electrical conductivity of the modified electrode. In comparison to the bare screen-printed electrode, the modified electrode showed a considerably higher performance and offered an excellent sensitivity along with a relatively fast analysis time.

An Electrochemistry and Computational Study at an Electrified Liquid-Liquid Interface for Studying Beta-Amyloid Aggregation.

Amphiphilic peptides, such as Aß amyloids, can adsorb at an interface between two immiscible electrolyte solutions (ITIES). Based on previous work (vide infra), a hydrophilic/hydrophobic interface is used as a simple biomimetic system for studying drug interactions. The ITIES provides a 2D interface to study ion-transfer processes associated with aggregation, as a function of Galvani potential difference. Here, the aggregation/complexation behaviour of Aß(1-42) is studied in the presence of Cu (II) ions, together with the effect of a multifunctional peptidomimetic inhibitor (P6). Cyclic and differential pulse voltammetry proved to be particularly sensitive to the detection of the complexation and aggregation of Aß(1-42), enabling estimations of changes in lipophilicity upon binding to Cu (II) and P6. At a 1:1 ratio of Cu (II):Aß(1-42), fresh samples showed a single DPV (Differential Pulse Voltammetry) peak half wave transfer potential (E1/2) at 0.40 V. Upon increasing the ratio of Cu (II) two-fold, fluctuations were observed in the DPVs, indicating aggregation. The approximate stoichiometry and binding properties of Aß(1-42) during complexation with Cu (II) were determined by performing a differential pulse voltammetry (DPV) standard addition method, which showed two binding regimes. A pKa of 8.1 was estimated, with a Cu:Aß1-42 ratio~1:1.7. Studies using molecular dynamics simulations of peptides at the ITIES show that Aß(1-42) strands interact through the formation of ß-sheet stabilised structures. In the absence of copper, binding/unbinding is dynamic, and interactions are relatively weak, leading to the observation of parallel and anti-parallel arrangements of ß-sheet stabilised aggregates. In the presence of copper ions, strong binding occurs between a copper ion and histidine residues on two peptides. This provides a convenient geometry for inducing favourable interactions between folded ß-sheet structures. Circular Dichroism spectroscopy (CD spectroscopy) was used to support the aggregation behaviour of the Aß(1-42) peptides following the addition of Cu (II) and P6 to the aqueous phase.

New Blatter-type radicals from a bench-stable carbene.

Stable benzotriazinyl radicals (Blatter's radicals) recently attracted considerable interest as building blocks for functional materials. The existing strategies to derivatize Blatter's radicals are limited, however, and synthetic routes are complex. Here, we report that an inexpensive, commercially available, analytical reagent Nitron undergoes a previously unrecognized transformation in wet acetonitrile in the presence of air to yield a new Blatter-type radical with an amide group replacing a phenyl at the C(3)-position. This one-pot reaction of Nitron provides access to a range of previously inaccessible triazinyl radicals with excellent benchtop stabilities. Mechanistic investigation suggests that the reaction starts with a hydrolytic cleavage of the triazole ring followed by oxidative cyclization. Several derivatives of Nitron were prepared and converted into Blatter-type radicals to test the synthetic value of the new reaction. These results significantly expand the scope of using functionalized benzotriazinyls as stable radical building blocks.

Emulsification at the Liquid/Liquid Interface: Effects of Potential, Electrolytes and Surfactants.

Emulsification of oils at liquid/liquid interfaces is of fundamental importance across a range of applications, including detergency. Adsorption and partitioning of the anionic surface active ions at the interface between two immiscible solutions is known to cause predictable chaos at the transfer potential region of the surfactant. In this work, the phenomenon that leads to the chaotic behaviour shown by sodium dodecylbenzene sulfonate (SDBS) at the water/1,2-dichloroethane interface is applied to commercial surfactants and aqueous/glyceryl trioleate interface. Electrochemical methods, electrocapillary curves, optical microscopy and conductivity measurements demonstrated that at 1.5 mm of SDBS, surfactants are adsorbed at the interface and assemble into micelles, leading to interfacial instability. As the concentration of the anionic surfactant was enhanced to 8 and 13.4 mm, the Marangoni effect and the interfacial emulsification became more prominent. The chaotic behaviour was found to be dependent on the surfactant concentration and the electrolytes present.

Platinum(II) complexes of N

A new family of platinum(II) complexes of the form PtL(n)SR have been prepared, where L(n) represents a cyclometalating, N^C^N-bound tridentate ligand and SR is a monodentate thiolate ligand. The complexes fall into two groups, those of PtL(1)SR where HL(1) = 1,3-bis(2-pyridyl)benzene, and those of PtL(2)SR, where HL(2) = methyl 3,5-bis(2-pyridyl)benzoate. Each group consists of five complexes, where R = CH3, C6H5, p-C6H4-CH3, p-C6H4-OMe, p-C6H4-NO2. These compounds, which are bright red, orange, or yellow solids, are formed readily upon treatment of PtL(n)Cl with the corresponding potassium thiolate KSR in solution at room temperature. The replacement of the chloride by the thiolate ligand is accompanied by profound changes in the photophysical properties. A broad, structureless, low-energy band appears in the absorption spectra, not present in the spectra of PtL(n)Cl. In the photoluminescence spectra, the characteristic, highly structured phosphorescence bands of PtL(n)Cl in the green region are replaced by a broad, structureless emission band in the red region. These new bands are assigned to a πS/dPt → π*N^C^N charge-transfer transition from the thiolate/platinum to the N^C^N ligand. This assignment is supported by electrochemical data and TD-DFT calculations and by the observation that the decreasing energies of the bands correlate with the electron-donating ability of the substituent, as do the increasing nonradiative decay rate constants, in line with the energy-gap law. However, the pair of nitro-substituted complexes do not fit the trends. Their properties, including much longer luminescence lifetimes, indicate that the lowest-energy excited state is localized predominantly on the arenethiolate ligand for these two complexes. Red-emitting thiolate adducts may be relevant to the use of PtL(n)Cl complexes in bioimaging, as revealed by the different distributions of emission intensity within live fibroplast cells doped with the parent complex, according to the region of the spectrum examined.

Modification of electrode surfaces by self-assembled monolayers of thiol-terminated oligo(phenyleneethynylene)s.

The wire-like properties of four S-(4-{2-[4-(2-phenylethynyl)phenyl]ethynyl}phenyl) thioacetate derivatives, PhC≡CC(6)H(4)C≡CC(6)H(4)SAc (1), H(2)NC(6)H(4)C≡CC(6)H(4)C≡CC(6)H(4)SAc (2), PhC≡CC(6)H(2)(OMe)(2)C≡CC(6)H(4)SAc (3) and AcSC(6)H(4)C≡CC(6)H(4)C≡CC(6)H(4)SAc (4) (Figure 1), all of which possess a high degree of conjugation along the oligo(phenyleneethynylene) (OPE) backbone, were investigated as self-assembled monolayers (SAMs) on gold and platinum electrodes by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The redox probe [Fe(CN)(6)](4)(-) was used in both the CV and impedance experiments. The results indicate that the thiolates derived from thioacetate-protected precursor molecules 1 and 2 form well-ordered monolayers on a gold electrode, whereas SAMs derived from 3 and 4 exhibit randomly distributed pinholes. The electron tunnelling resistance and fractional coverage of SAMs of all four compounds were examined using electron tunnelling theory. The analysis of the results reveal that the well-ordered SAMs of 1 and 2 exhibit higher charge-transfer resistance in comparison to the defect-ridden SAMs of 3 and 4. The additional steric bulk offered by the methoxy groups in 3 is likely to prevent efficient packing within the SAM, leading to a microelectrode behaviour, when assembled on a gold electrode surface. The protected dithiol derivative 4 probably binds to the surface through both terminal groups which prevents dense packing and leads to the formation of a monolayer with randomly distributed pinholes. Atomic force microscopy (AFM) was used to examine the morphology of the monolayers, and height images gave root-mean-square (RMS) roughness's which are in agreement with the proposed SAM structures.

Chiral acid selectivity displayed by PEDOT electropolymerised in the presence of chiral molecules.

Chiral conducting polymers prepared by electropolymerising PEDOT in the presence of chiral anions such as hyaluronic acid and anionic collagen or in a chiral nematic phase (hydroxypropyl cellulose, HPC) show excellent chiral acid recognition. This paper demonstrates the enantioselective recognition and transfer of protonated mandelic acid and protons using chiral PEDOTs. Discrimination between (R)-(-)- and (S)-(+)-mandelic acid was observed using cyclic voltammetry and square-wave voltammetry.

Electron transport in supported and tethered lipid bilayers modified with bioelectroactive molecules.

Tethered bilayer lipid membranes (tBLMs) are commonly used as model membranes. However, in biophysical studies, free-standing membranes ("black" lipid membranes) are more realistic models of cellular processes. In this article, we discuss the rates of electron transfer in both types of bilayer lipid membranes. These BLMs were then modified using two very important mitochondrial membrane-associated molecules: ubiquinone-10 (UQ10) and α-tocopherol (VitE). The electron transfer rates in the unmodified films were studied with three redox couples, Fe(CN)6(3-/4-), Ru(NH3)6(3+/2+), and NAD+/NADH, using electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The rate of electron transfer in the modified films was studied using the biologically relevant NAD+/NADH electroactive couple using the same methods. It is shown that when the BLMs are modified with only UQ10, it is possible to observe electron transfer. However, when the antioxidant VitE is added to the modification, the electron transfer provided by UQ10 is inhibited. Following initial studies using CV, a comparison of electron transfer theory and data was used to investigate this phenomenon in more detail, using EIS data. The standard rate constant caused by electron tunneling across the film, k(th)(0), depends on the value of ß used. Two different values of the potential independent electron tunneling coefficient, ß, were fitted, and it is shown that a ß value half of those usually reported in literature (refereed here as ß(app)) gives better agreement between the theory and the experimental results. The unmodified films present k(th)(0) values on the order of 10(-15) cm s(-1) when ß = 0.72 Å(-1) and k(th)(0) values on the order of 10(-9) cm s(-1) when ß(app) = 0.38 Å(-1). For the modified films, the values of k(th)(0) are on the order of 10(-15) cm s(-1) when ß = 0.72 Å(-1) and 10(-9) cm s(-1) for ß(app) = 0.38 Å(-1). The experimental electron transfer rate constant, k(app)(0), is on the order of 10(-8) cm s(-1) for unmodified and modified (with (i) UQ10, (ii) VitE, and (iii) UQ10 + VitE) films.

Chiral interactions of the drug propranolol and α1-acid-glycoprotein at a micro liquid-liquid interface.

The investigation of chiral interactions of drugs with plasma proteins is of fundamental importance for drug efficacy and toxicity studies. In this paper, we demonstrate a simple liquid-liquid interface procedure for investigating chiral interactions. Chiral discrimination of the enantiomers of a basic drug, propranolol, was achieved at a micro liquid-liquid interface, using α(1)-acid-glycoprotein (AGP) as a chiral acute phase plasma protein. When the protein is added to an aqueous phase containing the enantiomers of propranalol hydrochloride, the binding of (S)- and (R)-propranolol hydrochloride to the protein results in a decrease in the cyclic voltammetry (CV) and differential pulse voltammetry (DPV) current responses corresponding to the decrease in transfer of propranolol at an aqueous-1,2-dichloroethane interface. This decrease is a consequence of the complexation of the drug and the protein. The complex drug-protein does not transfer across the interface nor changes the transfer potential of the uncomplexed form of propranolol enantiomers. The bound concentration of propranolol enantiomers in the presence of AGP was found to be greater for (S)-propranolol than (R)-propranolol for solutions containing constant concentrations of AGP (50 µM). Scatchard analysis yielded association constants of 2.7 and 1.3 × 10(5) M(-1) for (S)- and (R)-propranolol, respectively.

A thermally actuated microgripper as an electrochemical sensor with the ability to manipulate single cells.

The design and fabrication of a thermally actuated polymeric microgripper incorporating a bare gold working electrode is described. Initial electrochemical tests confirm that the microgripper sensor works as an effective microelectrode, opening up exciting possibilities in single cell measurements.

Comparative electrochemical and impedance studies of self-assembled rigid-rod molecular wires and alkanethiols on gold substrates.

A study of the charge transfer and self-assembly characteristics of two new rigid-rod molecular wires 1 and 2 assembled on polycrystalline gold electrodes was carried out using electrochemical impedance spectroscopy and cyclic voltammetry. This class of wires have precisely controlled (ca. 1.5-2.5 nm) lengths of π-conjugation, with extended HOMO and LUMO wavefunctions. While rotations can occur around the C-C single bonds, the molecules cannot isomerise or fold due to their rigid backbone structures. The behaviour of these wires was compared with SAMs of heptanethiol (HPT) and dodecanethiol (DDT). It was found that SAMs of 1, which bears flexible hexyloxy sidechains, had randomly distributed pinholes which show microelectrode behaviour even when diluted with DDT. SAMs of 2, which do not have any sidechains, were well-organised at open-circuit potentials enabling evaluation of electron transfer kinetics assuming an average film thickness. However, impedance studies show that deviations from open circuit potentials resulted in an exponential decrease in charge transfer resistance, whereas capacitance remained constant, possibly attributable to conformational changes of the SAM. The syntheses and characterisation of the molecules is described.

Modification of the chiral selectivity of D-glucose oxidase and L-lactate oxidase in a collagen matrix.

A chiral, biocompatible, conducting material comprising collagen I, silica gel and ferrocene was found to modify and invert, the chiral selectivity of D-glucose oxidase and L-lactate oxidase. This result may have implications as collagen matrices are widely used in drug delivery, biosensing and tissue engineering. Investigations were carried out using electrochemical methods, FTIR spectroscopy and circular dichroism.

Chiral detection at a liquid-liquid interface.

Chiral ion transfer and detection at a liquid-liquid interface using chiral stationary phases such as cyclodextrins; could lead to alternative methods of chiral detection and separation.

Towards multifunctional microelectrode arrays.

A single microelectrode array platform, divided into groups of microelectrodes with flexible multiplexing can enable a 'defective' cluster of microelectrodes to be readily identified, by comparing normalised currents with the numbers of microelectrodes in the groups of arrays. This generic design principle may be extended for integrating multiple analyte sensing in a single microarray platform. The calibration of the microelectrode arrays, using white light interferometry and electrochemistry is described. The application for multianalyte detection is discussed.

Enantioselectivity of potentiometric sensors with application of different mechanisms of chiral discrimination.

In the recent years, numerous successful applications of various chiral selectors in high performance separation methods have generated an increasing interest in the application of some of these compounds as electroactive species in potentiometric sensors. The objective of this work was to examine the enantioselectivy of several different sensors employing substituted cyclodextrins, example antibiotic teicoplanin and electrodeposited conductive polymers for various chiral analytes. Varying degrees of enantioselectivity were found for the ion-selective electrodes examined, depending on the chiral selector used and the target analyte.

Mutations in the FAD binding domain cause stress-induced misoxidation of the endoplasmic reticulum oxidoreductase Ero1beta.

Disulfide bond catalysis is an essential component of protein biogenesis in the secretory pathway, from yeast through to man. In the endoplasmic reticulum (ER), protein-disulfide isomerase (PDI) catalyzes the oxidation and isomerization of disulfide bonds and is re-oxidized by an endoplasmic reticulum oxidoreductase (ERO). The elucidation of ERO function was greatly aided by the genetic analysis of two ero mutants, whose impairment results from point mutations in the FAD binding domain of the Ero protein. The ero1-1 and ero1-2 yeast strains have conditional and dithiothreitol-sensitive phenotypes, but the effects of the mutations on the behavior of Ero proteins has not been reported. Here, we show that these Gly to Ser and His to Tyr mutations do not prevent the dimerization of Ero1beta or the non-covalent interaction of Ero1beta with PDI. However, the Gly to Ser mutation abolishes disulfide-dependent PDI-Ero1beta heterodimers. Both the Gly to Ser and His to Tyr mutations make Ero1beta susceptible to misoxidation and aggregation, particularly during a temperature or redox stress. We conclude that the Ero FAD binding domain is critical for conformational stability, allowing Ero proteins to withstand stress conditions that cause client proteins to misfold.

Recommendation for measuring and reporting chloride by ISEs in undiluted serum, plasma or blood.

The proposed recommendation for measuring and reporting chloride in undiluted plasma or blood by ion-selective electrodes (ISEs) will provide results that are identical to chloride concentrations measured by coulometry for standardized normal plasma or blood samples. It is applicable to all current ISEs dedicated to chloride measurement in undiluted samples that meet the requirements. However, in samples with reduced water concentration, results by coulometry are lower than by ion-selective electrode due to volume displacement. The quantity measured by this standardized ISE procedure is called the ionized chloride concentration. It may be clinically more relevant than the chloride concentration as determined by coulometry, photometry or by ISE after dilution of the sample.

An introduction to thiol redox proteins in the endoplasmic reticulum and a review of current electrochemical methods of detection of thiols.

This aim of this paper is to expound the complexity of thiol redox systems in the endoplasmic reticulum of eukaryotic cells to the electroanalytical community. A summary of the state of the art in electrochemical methods for detection of thiols gives an insight into the challenges that need to be addressed to bridge the disparity between current analytical techniques and applications in a 'real' biological scenario.

Modular assembly of a preorganised, ditopic receptor for dicarboxylates.

Two types of calix[4]arene derived hosts for anions with, respectively, 1,3-alternate and cone conformations have been prepared; the 1,3-alternate system binds dicarboxylate anions in a ditopic manner while the cone compounds are deprotonated by carboxylates.

Individually addressable recessed gold microelectrode arrays with monolayers of thio-cyclodextrin nanocavities.

Four, individually addressable 30 microm diameter, e-beam deposited, gold microelectrodes recessed by 6 microm were suitably spaced on a single substrate to avoid diffusional overlap between each microelectrode. The single substrate device was functionalised with thiolated alpha-, beta-, and gamma-cyclodextrin nanocavities without spacer groups to ensure close proximity of the cavities to the electrode surface. The microelectrodes were assessed in two stages. The e-beam deposited micron sized electrodes were characterized using models for recessed and inlaid microdisk electrodes. Subsequently, each individually addressable, atomically flat, micro-patterned gold electrode with thiolated CD ensembles was treated as a nanoporous electrode assembly. Theoretical and experimental results were compared using cyclic voltammetry. Atomic force microscopy was also used to characterise the modified microelectrodes. Comparisons were made with thiolated CDs deposited on macroelectrodes. This is the first report of the behaviour of immobilized CD nanocavities ensembles on atomically flat gold microelectrodes.