Reversed phase behaviour of high molecular mass polystyrenes in ethyl acetate solvent mixtures

Chromatographia - The reversed phase behaviour of high molecular mass polystyrenes was investigated on C18 bonded phase columns with ethyl acetate-methanol and ethyl acetateacetonitrile mobile...     

Crystallization of Arthrobacter sp. strain 1C N-(1-D-carboxyethyl)-L-norvaline dehydrogenase and its complex with NAD+

The novel NAD+-linked opine dehydrogenase from a soil isolate Arthrobacter sp. strain 1C belongs to an enzyme superfamily whose members exhibit quite diverse substrate specificites. Crystals of this opine dehydrogenase, obtained in the presence or absence of co-factor and substrates, have been shown to diffract to beyond 1.8 ? resolution. X-ray precession photographs have established that the crystals belong to space group P21212, with cell parameters a = 104.9, b = 80.0, c = 45.5 ? and a single subunit in the asymmetric unit. The elucidation of the three-dimensional structure of this enzyme will provide a structural framework for this novel class of dehydrogenases to enable a comparison to be made with other enzyme families and also as the basis for mutagenesis experiments directed towards the production of natural and synthetic opine-type compounds containing two chiral centres.     

Capacity factors and peak widths in the gradient elution reversed phase separation of high molar mass polystyrenes

Summary A simplex method for determining the constantsS andk _o from the equation lnk’=lnk _o−S ϕ (was developed and applied to the reversed phase separation of high molar mass polystyrenes using gradients of any curvature. Experimental retention times described using the equation had a standard deviation of 1.1%. The inclusion of a quadratic term in the equation was found to be unwarranted. BothS and lnk _o varied linearly with ln molar mass. The logarithm of peak width of an eluting polystyrene peak was found to be a linear function of the mobile phase composition at elution. The slope was equal toS.     

Enhanced fluorescence from arrays of nanoholes in a gold film

Arrays of sub-wavelength holes (nanoholes) in gold films were used as a substrate for enhanced fluorescence spectroscopy. Seven arrays of nanoholes with distinct periodicities (distances between the holes) were fabricated. The arrays were then spin-coated with polystyrene films containing different concentrations of the fluorescent dye oxazine 720. The dye was excited via resonant extraordinary transmission of the laser source through the nanoholes. Enhanced fluorescence was observed when the geometric characteristics of the arrays allowed for an enhancement in the transmitted excitation. This enhancement occurred via surface plasmon excitation by the laser and a consequential increase in the local electromagnetic field in a sub-wavelength region at the metal-film interface. It was demonstrated that the sensitivity of the fluorescence measurement (change in signal vs change in dye concentration in the polymer film) is significantly larger at the surface plasmon resonance conditions than that obtained from equivalent films on glass substrates. Enhancement factors for the fluorescence emission were calculated for each array, with a maximum enhancement of close to 2 orders of magnitude as compared to the emission of films on glass. The results presented here indicate that arrays of nanoholes are interesting substrates for the development of fluorescence sensors based on surface plasmon resonance, as they provide a platform that allows both spatial confinement and enhancement of excitation light. Moreover, the collinear characteristics of the present optical setup, due to the resonant extraordinary transmission through the nanohole arrays, are more conducive to miniaturization and chip integration than more traditional experimental geometries.     

Mediated glucose enzyme electrodes by cross-linking films of osmium redox complexes and glucose oxidase on electrodes

Here, we report on a novel, versatile approach for the preparation of mediated enzyme electrodes, demonstrated using cross-linked films of glucose oxidase and a range of functionalised osmium complexes on graphite electrodes. Response of enzyme electrodes are optimised by evaluation of glucose response as a function of variation in ratios of [Os(2,2′-bipyridine)₂(4-aminomethyl pyridine)Cl]⁺ redox mediator, polyallylamine support and glucose oxidase enzyme cross-linked using a di-epoxide reagent in films on graphite. Lowering of the redox potential required to mediate glucose oxidation is achieved by synthesis of complexes using (4,4′-dimethyl-2,2′-bipyridine) or (4,4′-dimethoxy-2,2′-bipyridine) as a ligand instead of (2,2′-bipyridine). Enzyme electrodes prepared using the complexes based on dimethoxy- or dimethyl-substituted bipyridines provide glucose oxidation current densities of 30 and 70 μA?cm^?2 at 0.2 and 0.35 V applied potential compared to 120 μA?cm^?2 at 0.45 V for the initial enzyme electrode, under pseudo-physiological conditions in 5 mM glucose, with stability of signals proving inadequate for long-term operation. Current output and stability may be improved by selection of alternate anchoring and cross-linking methodology, to provide enzyme electrodes capable for application to long-term glucose biosensors and anodes in enzymatic fuel cells.

Studies on the metabolism and the detectability of 4-methyl-amphetamine and its isomers 2-methyl-amphetamine and 3-methyl-amphetamine in rat urine using GC-MS and LC-(high-resolution)-MS n

4-Methyl-amphetamine (1-(4-methylphenyl)propane-2-amine; 4-MA) and its isomers 2-methyl-amphetamine (2-MA) and 3-methyl-amphetamine (3-MA) belong to the group of amphetamine-type stimulants and of new psychoactive substances. Several studies showed similar potencies in releasing noradrenalin and dopamine, but higher potencies in releasing serotonin than amphetamine. In March 2013, the EU Council decided on an EU-wide control based on the European Monitoring Centre for Drugs and Drug Addiction risk assessment report documenting that 4-MA was sold as amphetamine on the illicit market and detected in several fatal cases. Therefore, 4-MA and its isomers should be covered by drug testing in clinical and forensic toxicology. The aims of the presented work were to study the metabolism and detectability of each isomer in urine samples. For metabolism studies, rat urine samples were isolated by solid-phase extraction without and after enzymatic cleavage of conjugates. The phase I metabolites were separated and identified after acetylation by gas chromatography–mass spectrometry (GC-MS) and/or liquid chromatography–high resolution-linear ion trap mass spectrometry (LC-HR-MS ⁿ ) and the phase II metabolites by LC-HR-MS ⁿ . From the identified phase I and II metabolites, the following main metabolic pathways were deduced: aromatic hydroxylation, hydroxylation of the phenylmethyl group followed by oxidation to the corresponding carboxylic acid, hydroxylation of the side chain, and glucuronidation and/or sulfation of the hydroxy and carboxy groups. CYP2D6 was involved in the aromatic hydroxylation. Finally, the intake of a commonly used dose of the MAs could be confirmed in rat urine using the authors’ GC-MS and the LC-MS ⁿ standard urine screening approaches. Differentiation of the isomers to confirm the intake of a specific isomer was possible with an additional workup in rat urine.     

Designing of an Intensification Process for Biosynthesis and Recovery of Menaquinone-7

A nutritional food rich in menaquinone-7 has a potential in preventing osteoporosis and cardiovascular diseases. The static fermentation of Bacillus subtilis natto is widely regarded as an optimum process for menaquinone-7 production. The major issues for the bulk production of menaquinone-7 are the low fermentation yield, biofilm formation and the use of organic solvents for the vitamin extraction. In this study, we demonstrate that the dynamic fermentation involving high stirring and aeration rates enhances the yield of fermentation process significantly compared to static system. The menaquinone-7 concentration of 226 mg/L was produced at 1,000 rpm, 5 vvm, 40 °C after 5 days of fermentation. This concentration is 70-fold higher than commercially available food products such as natto. Additionally, it was found that more than 80 % of menaquinone-7 was recovered in situ in the vegetable oil that was gradually added to the system as an anti-foaming agent. The intensification process developed in this study has a capacity to produce an oil rich in menaquinone-7 in one step and eliminate the use of organic solvents for recovery of this compound. This oil can, therefore, be used for the preparation of broad range of supplementary and dietary food products rich in menaquinone-7 to reduce the risk of osteoporotic fractures and cardiovascular diseases.     

Experimental setup and first measurement of DNA damage induced along and around an antiproton beam

Radiotherapy employs ionizing radiation to induce lethal DNA lesions in cancer cells while minimizing damage to healthy tissues. Due to their pattern of energy deposition, better therapeutic outcomes can, in theory, be achieved with ions compared to photons. Antiprotons have been proposed to offer a further enhancement due to their annihilation at the end of the path. The work presented here aimed to establish and validate an experimental procedure for the quantification of plasmid and genomic DNA damage resulting from antiproton exposure. Immunocytochemistry was used to assess DNA damage in directly and indirectly exposed human fibroblasts irradiated in both plateau and Bragg peak regions of a 126 MeV antiproton beam at CERN. Cells were stained post irradiation with an anti-γ-H2AX antibody. Quantification of the γ-H2AX foci-dose relationship is consistent with a linear increase in the Bragg peak region. A qualitative analysis of the foci detected in the Bragg peak and plateau region indicates significant differences highlighting the different severity of DNA lesions produced along the particle path. Irradiation of desalted plasmid DNA with 5 Gy antiprotons at the Bragg peak resulted in a significant portion of linear plasmid in the resultant solution.     

Low-energy behavior of the 3He(α, γ)7Be cross section

Cross sections for the ³He(α, γ)⁷Be reaction have been measured at several energies from E_c.m. = 165 to 1169 keV by counting prompt γ-rays from a windowless, differentially pumped, recirculating, ³He gas target. The cross-section factor S₃₄(E_c.m.) and branching ratio γ₁/γ₀ were determined at each energy. Cross sections were also measured at E_c.m. = 947 and 1255 keV by counting the γ-rays from the ⁷Be produced in a ³He gas cell with a Ni entrance foil. Combining the results of these two independent experiments yields a zero-energy intercept for the cross-section factor of S₃₄(0) = 0.53 ± 0.03 keV · b. The relationship between these measurements and several theoretical calculations, and the import of the extrapolated cross section for the solar-neutrino problem are discussed.