Preparation of highly accessible mordenite coatings on ceramic monoliths at loadings exceeding 50% by weight

A mordenite layer with a high accessibility has been synthesised on cordierite monolith supports; substantial loadings of mordenite were achieved (above 50 wt%) under the synthesis conditions used.     

On the Possibility of Uphill Intramolecular Electron Transfer in Multicopper Oxidases: Electrochemical and Quantum Chemical Study of Bilirubin Oxidase

The catalytic cycle of multicopper oxidases (MCOs) involves intramolecular electron transfer (IET) from the Cu‐T1 copper ion, which is the primary site of the one‐electron oxidations of the substrate, to the trinuclear copper cluster (TNC), which is the site of the four‐electron reduction of dioxygen to water. In this study we report a detailed characterization of the kinetic and electrochemical properties of bilirubin oxidase (BOx) – a member of the MCO family. The experimental results strongly indicate that under certain conditions, e.g. in alkaline solutions, the IET can be the rate‐limiting step in the BOx catalytic cycle. The data also suggest that one of the catalytically relevant intermediates (most likely characterized by an intermediate oxidation state of the TNC) formed during the catalytic cycle of BOx has a redox potential close to 0.4 V, indicating an uphill IET process from the T1 copper site (0.7 V) to the Cu‐T23. These suggestions are supported by calculations of the IET rate, based on the experimentally observed Gibbs free energy change and theoretical estimates of reorganization energy obtained by combined quantum and molecular mechanical (QM/MM) calculations.     

Effects of sample preservation and storage on mercury speciation in natural stream water

Despite an increasing focus on low level methods for determination of mercury species in water over the last decades, few studies have paid attention to direct effects of different sample preparation methods (i.e. preservation techniques) on natural freshwater samples. In this study we show how different preservation techniques give significantly different concentrations of total and methylmercury in freshwaters (9 and 14% on average, respectively). Natural stream samples from a forested lake catchment were studied. Mean stream sample concentrations of total (3.6 ng/L) and methylmercury (0.06 ng/L) reflect levels typical for pristine humic boreal catchments. The main reason for the observed average differences in total and methylmercury concentrations is the use of one instead of two sample bottles and timing of sample acidification, respectively.     

Quantitative detection of well-based DNA array using switchable lanthanide luminescence

In this report a novel wash-free method for multiplexed DNA detection is demonstrated employing target specific probe pairs and switchable lanthanide luminescence technology on a solid-phase array. Four oligonucleotide capture probes, conjugated at 3′ to non-luminescent lanthanide ion carrier chelate, were immobilized as a small array on the bottom of a microtiter plate well onto which a mix of corresponding detection probes, conjugated at 5′ to a light absorbing antenna ligand, were added. In the presence of complementary target nucleic acid both the spotted capture probe and the liquid-phase detection probe hybridize adjacently on the target. Consequently the two non-luminescent label molecules self-assemble and form a luminescent mixed lanthanide chelate complex. Lanthanide luminescence is thereafter measured without a wash step from the spots by scanning in time-resolved mode. The homogeneous solid-phase array-based method resulted in quantitative detection of synthetic target oligonucleotides with 0.32 nM and 0.60 nM detection limits in a single target and multiplexed assay, respectively, corresponding to 3× SD of the background. Also qualitative detection of PCR-amplified target from Escherichia coli is described.     

DNA-controlled assembly of soft nanoparticles

Immobilization of DNA (encoding) on solid nanoparticles requires surface chemistry, which is well established for gold surfaces but often tedious and not generally applicable for many other inorganic surface materials. While substantial effort has been devoted to expanding surface chemistry techniques for solid nanoparticles, considerably less attention has been given to the development of noncovalent attachment of DNA to soft nanoparticles, like liposomes. Here we report a DNA-controlled assembly of liposomes in solution and on solid supported membranes, this process displays remarkably sharp thermal transitions from an assembled to a disassembled state, allowing application of DNA-controlled liposome assembly for the detection of polynucleotides (e.g., DNA) with single mismatch discrimination power. The method is based on a single DNA strand (contains two lipid membrane anchors), which is able to noncovalently attach to a liposome surface. This design enables detection of biological polynucleotide targets as the complementary strand can be unmodified DNA and RNA strands.     

Monitoring of Saccharomyces cerevisiae cell proliferation on thiol-modified planar gold microelectrodes using impedance spectroscopy

An impedance spectroscopic study of the interaction between thiol-modified Au electrodes and Saccharomyces cerevisiae of strain EBY44 revealed that the cells formed an integral part of the interface, modulating the capacitive properties until a complete monolayer was obtained, whereas the charge transfer resistance ( R ct) to the redox process of [Fe(CN)6] 3-/4- showed a linear relationship to the number of cells even beyond the monolayer coverage. R ct showed strong pH dependence upon increasing the pH of the utilized buffer to 7.2. Upon addition of S. cerevisiae cells at pH 7.2, the obtained value of R ct showed over 560% increase with respect to the value obtained on the same thiol-modified electrode without cells. It was demonstrated that real-time monitoring of S. cerevisiae proliferation, with frequency-normalized imaginary admittance (real capacitance) as the indicator, was possible using a miniaturized culture system, ECIS Cultureware, with integrated planar cysteamine-modified Au microelectrodes. A monolayer coverage was reached after 20-28 h of cultivation, observed as an approximately 15% decrease in the real capacitance of the system.     

Adsorption and activity of Thermomyces lanuginosus lipase on hydrophobic and hydrophilic surfaces measured with dual polarization interferometry (DPI) and confocal microscopy

The adsorption and activity of Thermomyces lanuginosus lipase (TLL) was measured with dual polarization interferometry (DPI) and confocal microscopy at a hydrophilic and hydrophobic surface. In the adsorption isotherms, it was evident that TLL both had higher affinity for the hydrophobic surface and adsorbed to a higher adsorbed amount (1.90 mg/m²) compared to the hydrophilic surface (1.40–1.50 mg/m²). The thickness of the adsorbed layer was constant (3.5 nm) on both surfaces at an adsorbed amount >1.0 mg/m², but decreased on the hydrophilic surface at lower surface coverage, which might be explained by partially unfolding of the TLL structure. However, a linear dependence of the refractive index of the adsorbed layer on adsorbed amount of TLL on C18 surfaces indicated that the structure of TLL was similar at low and high surface coverage. The activity of adsorbed TLL was measured towards carboxyfluorescein diacetate (CFDA) in solution, which upon lipase activity formed a fluorescent product. The surface fluorescence intensity increase was measured in a confocal microscope as a function of time after lipase adsorption. It was evident that TLL was more active on the hydrophilic surface, which suggested that a larger fraction of adsorbed TLL molecules were oriented with the active site facing the solution compared to the hydrophobic surface. Moreover, most of the activity remained when the TLL surface coverage decreased. Earlier reports on TLL surface mobility on the same surfaces have found that the lateral diffusion was highest on hydrophilic surfaces and at low surface coverage of TLL. Hence, a high lateral mobility might lead to a longer exposure time of the active site towards solution, thereby increasing the activity against a water-soluble substrate.     

A membrane-, mediator-, cofactor-less glucose/oxygen biofuel cell

We report the fabrication and characterisation of a non-compartmentalised, mediator and cofactor free glucose-oxygen biofuel cell based on adsorbed enzymes exhibiting direct bioelectrocatalysis, viz. cellobiose dehydrogenase from Dichomera saubinetii and laccase from Trametes hirsuta as the anodic and cathodic bioelements, respectively, with the following characteristics: an open-circuit voltage of 0.73 V; a maximum power density of 5 microW cm(-2) at 0.5 V of the cell voltage and an estimated half-life of > 38 h in air-saturated 0.1 M citrate-phosphate buffer, pH 4.5 containing 5 mM glucose.