Multi-detection method for five common microalgal toxins based on the use of microspheres coupled to a flow-cytometry system

Freshwater and brackish microalgal toxins, such as microcystins, cylindrospermopsins, paralytic toxins, anatoxins or other neurotoxins are produced during the overgrowth of certain phytoplankton and benthic cyanobacteria, which includes either prokaryotic or eukaryotic microalgae. Although, further studies are necessary to define the biological role of these toxins, at least some of them are known to be poisonous to humans and wildlife due to their occurrence in these aquatic systems. The World Health Organization (WHO) has established as provisional recommended limit 1 μg of microcystin-LR per liter of drinking water. In this work we present a microsphere-based multi-detection method for five classes of freshwater and brackish toxins: microcystin-LR (MC-LR), cylindrospermopsin (CYN), anatoxin-a (ANA-a), saxitoxin (STX) and domoic acid (DA). Five inhibition assays were developed using different binding proteins and microsphere classes coupled to a flow-cytometry Luminex system. Then, assays were combined in one method for the simultaneous detection of the toxins. The IC₅₀'s using this method were 1.9 ± 0.1 μg L⁻¹ MC-LR, 1.3 ± 0.1 μg L⁻¹ CYN, 61 ± 4 μg L⁻¹ ANA-a, 5.4 ± 0.4 μg L⁻¹ STX and 4.9 ± 0.9 μg L⁻¹ DA. Lyophilized cyanobacterial culture samples were extracted using a simple procedure and analyzed by the Luminex method and by UPLC–IT-TOF-MS. Similar quantification was obtained by both methods for all toxins except for ANA-a, whereby the estimated content was lower when using UPLC–IT-TOF-MS. Therefore, this newly developed multiplexed detection method provides a rapid, simple, semi-quantitative screening tool for the simultaneous detection of five environmentally important freshwater and brackish toxins, in buffer and cyanobacterial extracts.

Numerical Method for Cost-Weight Optimisation of Stringer-Skin Panels

The need to integrate cost into the early product definition process as an engineering parameter is addressed. The application studied is a fuselage panel that is typical for commercial transport regional jets. Consequently, a semi-empirical numerical analysis using reference data was coupled to model the structural integrity of thin-walled structures with regard to material failure and buckling: skin, stringer, flexural, and interrivet. The optimization process focuses on direct operating cost (DOC) as a function of acquisition cost and fuel burn. It was found that the ratio of acquisition cost to fuel burn was typically 4:3 and that there was a 10% improvement in the DOC for the minimal DOC condition over the minimal weight condition because of the manufacturing cost saving from having a reduced number of larger-area stringers and a slightly thicker skin than that preferred by the minimal weight condition. Also note that the minimal manufacturing cost condition was slightly better than the minimal weight condition, which highlights the key finding: The traditional minimal weight condition is a dated and suboptimal approach to airframe structural design.

Finite-Element Time-Step Coupled Generator, Load, AVR and Brushless Exciter Modelling

This paper presented results from a details and comprehensive simulation using finite element method of the practical operation of an electrical machine. The results it displayed have been used in practice to design more efficient equipment.

Coupled heat and moisture transfer in multi-layer building materials

A dynamic mathematical model for simulating the coupled heat and moisture migration through multilayer porous building materials was proposed. Vapor content and temperature were chosen as the principal driving potentials. The discretization of the governing equations was done by the finite difference approach. A new experimental set-up was also developed in this study. The evolution of transient temperature and moisture distributions inside specimens were measured. The method for determining the temperature gradient coefficient was also presented. The moisture diffusion coefficient, temperature gradient coefficient, sorption–desorption isotherms were experimentally evaluated for some building materials (sandstone and lime-cement mortar). The model was validated by comparing with the experimental data with good agreement. Another advantage of the method lies in the fact that the required transport properties for predicting the non-isothermal moisture flow only contain the vapor diffusion coefficient and temperature gradient coefficient. They are relatively simple, and can be easily determined.

Development of a simple gel permeation clean-up procedure coupled to a rapid disequilibrium enzyme-linked immunosorbent assay (ELISA) for the detection of Sudan I dye in spices and sauces

Sudan dyes have been found to be added to chilli and chilli products for illegal colour enhancement purposes. Due to the possible carcinogenic effect, they are not authorized to be used in food in the European Union or the USA. However, over the last few years, many products imported from Asian and African countries have been reported via the Rapid Alert System for Food and Feed in the European Union to be contaminated with these dyes. In order to provide fast screening method for the detection of Sudan I (SI), which is the most widely abused member of Sudan dyes family, a unique (20 min without sample preparation) direct disequilibrium enzyme-linked immunosorbent assay (ELISA) was developed. The assay was based on polyclonal antibodies highly specific to SI. A novel, simple gel permeation chromatography clean-up method was developed to purify extracts from matrices containing high amounts of fat and natural pigments, without the need for a large dilution of the sample. The assay was validated according to the Commission Decision 2002/657/EC criteria. The detection capability was determined to be 15 ng g(-1) in sauces and 50 ng g(-1) in spices. The recoveries found ranged from 81% to 116% and inter- and intra-assay coefficients of variation from 6% to 20%. The assay was used to screen a range of products (85 samples) collected from different retail sources within and outside the European Union. Three samples were found to contain high amounts (1,649, 722 and 1,461 ng g(-1)) of SI by ELISA. These results were confirmed by liquid chromatography-tandem mass spectrometry method. The innovative procedure allows for the fast, sensitive and high throughput screening of different foodstuffs for the presence of the illegal colorant SI.

Apparatus and method for adjusting filter frequency in relation to sampling frequency

A self-tuning filter is disclosed. The self-tuning filter includes a digital clocking signal and an input coupled to the digital clocking signal, whereby the input reads a value incident on the input when the digital clocking signal changes to a predetermined state. A clock-tunable filter is, furthermore, coupled to the digital clocking signal so that the frequency of the clock-tunable filter is adjusted in relation to a sampling frequency at which the digital clocking signal operates. The self-tuning filter may be applied to an input of a data acquisition unit and applied to an input having a variable sampling frequency. A method of controlling the frequency of a clock-tunable filter is also disclosed.

Development and validation of a method for the confirmation of halofuginone in chicken liver and eggs using electrospray tandem mass spectrometry

A method is described for the quantitative confirmation of halofuginone (HFG) residues in chicken liver and eggs. This method is based on LC coupled to positive ion electrospray MS-MS of the tissue extracts, prepared by trypsin digestion of the tissues followed by liquid-liquid extraction and final clean-up using Solid Phase Extraction (SPE). The [M+H](+) ion at m/z 416 is monitored along with four transitions at m/z 398, 138, 120 and 100. The method has been validated according to the draft EU criteria for the analysis of veterinary drug residues at 15, 30 and 45 mug kg (-1) in liver and 5, 15 and 50 mug kg (-1) in eggs. The new analytical limits, CCalpha and CCbeta were calculated for liver and were 35.4 and 43.6 mug kg (-1), respectively. (C) 2003 Elsevier Science B.V. All rights reserved.

Development and validation of a method for the confirmation of nicarbazin in chicken liver and eggs using LC-electrospray MS-MS according to the revised EU criteria for veterinary drug residue analysis

A method is described for the quantitative confirmation of 4,4'-dinitrocarbanilide (DNC), the marker residue for nicarbazin in chicken liver and eggs. The method is based on LC coupled to negative ion electrospray MS-MS of tissue extracts prepared by liquid-liquid extraction. The [M-H](-) ion at m/z 301 is monitored along with two transition ions at m/z 137 and 107 for DNC and the [M-H](-) ion at m/z 309 for the internal standard, d(8)-DNC. The method has been validated according to the new EU criteria for the analysis of veterinary drug residues at 100, 200 and 300 mug kg(-1) in liver and at 10, 30 and 100 mug kg(-1) in eggs. Difficulties concerning the application of the new analytical limits, namely the decision limit (CC) and the detection capability (CC) to the determination of DNC in both liver and eggs are discussed.

Unraveling the Structure and Function of G Protein-Coupled Receptors Through NMR Spectroscopy

G protein-coupled receptors (GPCRs) are a large superfamily of signaling proteins expressed on the plasma membrane. They are involved in a wide range of physiological processes and, therefore, are exploited as drug targets in a multitude of therapeutic areas. In this extent, knowledge of structural and functional properties of GPCRs may greatly facilitate rational design of modulator compounds. Solution and solid-state nuclear magnetic resonance (NMR) spectroscopy represents a powerful method to gather atomistic insights into protein structure and dynamics. In spite of the difficulties inherent the solution of the structure of membrane proteins through NMR, these methods have been successfully applied, sometimes in combination with molecular modeling, to the determination of the structure of GPCR fragments, the mapping of receptor-ligand interactions, and the study of the conformational changes associated with the activation of the receptors. In this review, we provide a summary of the NMR contributions to the study of the structure and function of GPCRs, also in light of the published crystal structures.

Investigation into mercury bound to biothiols:structural identification using ESI-ion-trap MS and introduction of a method for their HPLC separation with simultaneous detection by ICP-MS and ESI-MS

Mercury in plants or animal tissue is supposed to occur in the form of complexes formed with biologically relevant thiols (biothiols), rather than as free cation. We describe a technique for the separation and molecular identification of mercury and methylmercury complexes derived from their reactions with cysteine (Cys) and glutathione (GS): Hg(Cys)(2), Hg(GS)(2), MeHgCys, MeHgGS. Complexes were characterised by electrospray mass spectrometry (MS) equipped with an ion trap and the fragmentation pattern of MeHgCys was explained by using MP2 and B3LYP calculations, showing the importance of mercury-amine interactions in the gas phase. Chromatographic baseline separation was performed within 10 min with formic acid as the mobile phase on a reversed-phase column. Detection was done by online simultaneous coupling of ES-MS and inductively coupled plasma MS. When the mercury complexes were spiked in real samples (plant extracts), no perturbation of the separation and detection conditions was observed, suggesting that this method is capable of detecting mercury biothiol complexes in plants.

An in situ spatially resolved method to probe gas phase reactions through a fixed bed catalyst

A new method to spatially probe heterogeneous catalysed reactions within a packed bed of catalyst has been developed. The spatial resolution is achieved using a stationary perforated capillary coupled to a mass spectrometer while the catalyst bed is moved. The oxidation of CO promoted by H-2 over a Pd catalyst has been used to demonstrate the technique.

Prediction of Transonic LCOs using an Aeroelastic Harmonic Balance Method

This work proposes a novel approach to compute transonic Lim
it Cycle Oscillations using high fidelity analysis. CFD based Harmonic Balance methods have proven to be efficient tools to predict periodic phenomena. This paper’s contribution is to present a new methodology to determine the unknown frequency of oscillations, enabling HB methods to accurately capture Limit Cycle Oscillations (LCOs); this is achieved by defining a frequency updating procedure based on a coupled CFD/CSD Harmonic Balance formulation to find the LCO condition. A pitch/plunge aerofoil and delta wing aerodynamic and respective linear structural models are used to validate the new method against conventional time-domain simulations. Results show consistent agreement between the proposed and time-marching methods for both LCO amplitude and frequency, while producing at least one order of magnitude reduction in computational time.

A multiconfigurational time-dependent Hartree-Fock method for excited electronic states. II. Coulomb interaction effects in single conjugated polymer chains

Conjugated polymers have attracted considerable attention in the last few decades due to their potential for optoelectronic applications. A key step that needs optimisation is charge carrier separation following photoexcitation. To understand better the dynamics of the exciton prior to charge separation, we have performed simulations of the formation and dynamics of localised excitations in single conjugated polymer strands. We use a nonadiabatic molecular dynamics method which allows for the coupled evolution of the nuclear degrees of freedom and of multiconfigurational electronic wavefunctions. We show the relaxation of electron-hole pairs to form excitons and oppositely charged polaron pairs and discuss the modifications to the relaxation process predicted by the inclusion of the Coulomb interaction between the carriers. The issue of charge photogeneration in conjugated polymers in dilute solution is also addressed. (C) 2011 American Institute of Physics. [doi: 10.1063/1.3600404]