Interaction of self-assembled monolayers of DNA with electrons: HREELS and XPS studies

We present results from high-resolution electron energy loss spectroscopy (HREELS) and XPS studies of self-assembled monolayers of DNA. The monolayers are well-organized and display sharp vibrational peaks in the HREEL spectra. The electrons interact mainly with the backbone of the DNA. The XPS results indicate that, in most of the samples studied, the phosphates on the DNA are not charged.     

Removal of envelope protein-free retroviral vectors by anion-exchange chromatography to improve product quality

We have investigated the role of the retroviral lipid bilayer and envelope proteins in the adsorption of retroviral vectors (RVs) to a Fractogel DEAE matrix. Intact RVs and their degradation components (envelope protein-free vectors and solubilized vector components) were adsorbed to this matrix and eluted using a linear gradient. Envelope protein-free RVs (Env(-)) and soluble envelope proteins (gp70) eluted in a significantly lower range of conductivities than intact RVs (Env(+)) (13.7-30 mS/cm for Env(-) and gp70 proteins vs. 47-80 mS/cm for Env(+)). The zeta (zeta)-potential of Env(+) and Env(-) vectors was evaluated showing that envelope proteins define the pI of the viral particles (pI (Env(+)) < 2 versus 3 < pI (Env(-)) < 4) and that Env(+) and Env(-) vectors have similar zeta-potentials within pH 5 and 8. The results presented herein indicate that the adsorption of retroviral particles occurs through multi-point interaction of the envelope proteins with the cationic groups on the chromatographic matrix. The strength of this adsorption is thus dependent on the amount of envelope protein present in the viral lipid bilayer. In conclusion, AEXc enables the separation of gp70 proteins as well as envelope protein-free vectors constituting a significant improvement to the quality of retroviral preparations for gene therapy applications.     

Application of a polarity parameter model to the separation of fat-soluble vitamins by reversed-phase HPLC

The retention behavior of a series of fat-soluble vitamins has been established on the basis of a polarity retention model: log k = (log k)(0) + p (P(m) (N) - P(s) (N)), with p being the polarity of the solute, P(m) (N) the mobile phase polarity, and (log k)(0) and P(m) (N) two parameters for the characterization of the stationary phase. To estimate the p-values of solutes, two approaches have been considered. The first one is based on the application of a QSPR model, derived from the molecular structure of solutes and their log P(o/w), while in the second one, the p-values are obtained from several experimental measurements. The quality of prediction of both approaches has also been evaluated, with the second one giving more accurate results for the most lipophilic vitamins. This model allows establishing the best conditions to separate and determine simultaneously some fat-soluble vitamins in dairy foods.     

Determination of illicit and medicinal drugs and their metabolites in oral fluid and preserved oral fluid by liquid chromatography–tandem mass spectrometry

An LC-MS/MS method using 0.5 ml of oral fluid was developed for the determination of morphine, codeine, 6-monoacetylmorphine, methadone, amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxymethamphetamine, 3,4-methylenedioxy-N-ethylamphetamine, benzoylecgonine, cocaine, delta-9-tetrahydrocannabinol, zolpidem, zopiclone, alprazolam, clonazepam, oxazepam, nordiazepam, lorazepam, flunitrazepam, diazepam, diphenhydramine and amitriptyline. The method was fully validated in terms of linearity (the method was linear between 1–5 μg/L and 100–200 μg/L) recoveries (7.5–82.6%), within-day and between-day precisions and accuracies (CV and MRE, both <15%), limits of detection (0.5 μ g/L) and quantitation (the lowest point on the calibration curve), relative ion intensities, freeze-and-thaw stability and matrix effect. The method was applied to preserved oral fluid collected by a special commercial device, the StatSure Saliva Sampler™.     

Bohr’s strip for vector valued Dirichlet series

Bohr showed that the width of the strip (in the complex plane) on which a given Dirichlet series , converges uniformly but not absolutely, is at most 1/2, and Bohnenblust-Hille that this bound in general is optimal. We prove that for a given infinite dimensional Banach space Y the width of Bohr’s strip for a Dirichlet series with coefficients a _n in Y is bounded by 1 - 1/Cot (Y), where Cot (Y) denotes the optimal cotype of Y. This estimate even turns out to be optimal, and hence leads to a new characterization of cotype in terms of vector valued Dirichlet series. The first, second and third authors were supported by MEC and FEDER Project MTM2005-08210.     

Wavelet neural networks to resolve the overlapping signal in the voltammetric determination of phenolic compounds

Three phenolic compounds, i.e. phenol, catechol and 4-acetamidophenol, were simultaneously determined by voltammetric detection of its oxidation reaction at the surface of an epoxy-graphite transducer. Because of strong signal overlapping, Wavelet Neural Networks (WNN) were used in data treatment, in a combination of chemometrics and electrochemical sensors, already known as the electronic tongue concept. To facilitate calibration, a set of samples (concentration of each phenol ranging from 0.25 to 2.5mM) was prepared automatically by employing a Sequential Injection System. Phenolic compounds could be resolved with good prediction ability, showing correlation coefficients greater than 0.929 when the obtained values were compared with those expected for a set of samples not employed for training.     

Protein extraction from Phoenix dactylifera L. leaves, a recalcitrant material, for two-dimensional electrophoresis

This work was aimed at optimizing a protein extraction procedure for date palm (Phoenix dactylifera L.) leaves, a highly recalcitrant plant tissue for 2-DE. Five protein extraction protocols based on different protein precipitation agents (TCA/acetone vs. phenol (Ph) methods) and protein resolubilization methods (physical treatments, e.g., sonication, shaking and/or heating) were tested. Ph/SDS extraction with methanol/ammonium acetate precipitation, followed by DOC preincubation and TCA/acetone precipitation and, finally, solubilization by shaking in rehydration solution was found to be the best protein extraction method. We conclude that DOC with TCA/acetone precipitation step eliminates interfering compounds, thus allowing efficient resolubilization of date palm leaf proteins. This method could be appropriate for proteomic studies such as date palm colonization by entomopathogenic fungi.     

Comparison of stir bar sorptive extraction and membrane-assisted solvent extraction for the ultra-performance liquid chromatographic determination of oxazole fungicide residues in wines and juices

The present study compares two new sample preparation methods, stir bar sorptive extraction (SBSE) and membrane-assisted solvent extraction (MASE) coupled to the novel technique of ultra-performance liquid chromatography (UPLC) for the sensitive, selective and solvent-free determination of six oxazole fungicide residues (hymexazol, drazoxolon, vinclozolin, chlozolinate, oxadixyl and famoxadone) in wine and juices. The analytes were separated on a rapid resolution C(18) column (50 mm x 4.6 mm, I.D., 1.8 microm) thermostated at 50 degrees C with isocratic elution using a 50/50 (v/v) water/acetonitrile (ACN) mobile phase at a flow-rate of 1 mL min(-1) and detected by diode-array detection (DAD). The UPLC method rapidly separates the fungicides (7 min). The best results as regards sensitivity, repeatability and analyte recovery were obtained using SBSE with a polydimethylsiloxane (PDMS) twister, at 60 degrees C for 30 min with stirring at 1700 rpm in the presence of a 0.1M acetate/acetic acid buffer (pH 5) and 20% (m/v) sodium chloride. Liquid desorption was performed with 100 microL of a 80/20 (v/v) ACN/water solution in a desorption time of 15 min. With the PDMS polymer, an apolar phase, hymexazol and oxadixyl were not extracted. Consequently, the SBSE procedure can only be applied to the other four fungicides. Detection limits ranged from 0.05 to 2.5 microgL(-1) at a signal to noise ratio of 3, depending on the compound. Recoveries obtained for spiked samples were satisfactory (83-113%) for all compounds. The proposed method was successfully applied to the analysis of different samples, residues of chlozolinate and drazoxolon being found in samples of red wine and grape juice, respectively.