Stereo-specific synthesis of hydroanthracene-dicarboximides

Compound 3, N-((1S)-1-cyclohexylethyl)-9,10-dihydro-9,10-ethanoanthracene-(11S,12S)-dicarboximide-1,2,3,4-octahydro, was obtained by ruthenium-assisted hydrogenation of the hydroanthracene-dicarboximide 2 under mild conditions (3 bar H₂ and room temperature). In contrast to other related compounds, dicarboximides 2 and 3 were stereo-selectively obtained, confirmed by both solid state (X-ray diffraction) and solution (NMR). This selectivity denoted a hindered rotation around the N-CH axis together with the aromatic hydrogen bond acceptor behaviour of the hydroanthracene skeleton towards a methylene of the cyclohexyl group of the imide moiety. In addition, the nature of the metallic species involved in the hydrogenation process was also investigated.     

C(5)N(-) anion and new carbenic isomers of cyanodiacetylene: a matrix isolation IR study

Products of the vacuum-UV photolysis of cyanodiacetylene (HC(5)N) in solid argon -- the anion C(5)N(-), imine HNC(5), and the branched carbene C(4)(H)CN -- have been identified by IR absorption spectroscopy, in addition to the already discovered isonitrile HC(4)NC. Spectral assignments were assisted by deuterium substitution experiments, by BD(T) calculations, and by the results of a recent density functional theory study.     

Field-amplified sample stacking for the detection of chemical warfare agent degradation products in low-conductivity matrices by capillary electrophoresis-mass spectrometry

Preconcentration of chemical warfare agent degradation products (alkylphosphonic acids and alkyl alkylphosphonic acids) in low-conductivity matrices (purified water, tap water and local river water) by field-amplified sample stacking (FASS) was developed for capillary electrophoresis (CE) coupled to ion trap mass spectrometry. FASS was performed by adding a mixture of HCOONH(4) and NH(4)OH in appropriate concentrations to the sample. This allowed to control the conductivity and the pH of the sample in order to obtain FASS performances that are independent of analyte concentration. The influence of different parameters on FASS (sample to background electrolyte (BGE) conductivity ratio, injection volume and concentration of BGE) was studied to determine the optimal conditions and was rationalized by using the theoretical model developed by Burgi and Chien. A good correlation was obtained between the bulk electroosmotic velocity predicted by this model and the experimental value deduced from the migration time of the electroosmotic flow marker detected by mass spectrometry (MS). This newly developed method was successfully applied to the analysis of tap water and local river water fortified with the analytes and provided a 10-fold sensitivity enhancement in comparison to the signal obtained without preconcentration procedure. The quite satisfactory repeatability and linearity for peak areas obtained in the 0.5-5 microg mL(-1) concentration range allow quantitative analysis to be implemented. Limits of detection of 0.25-0.5 microg mL(-1) for the alkyl alkylphosphonic acids and of 0.35-5 microg mL(-1) for the alkylphosphonic acids were reached in tap water and river water.     

Determination of galactose and mannose residues in natural galactomannans using a fast and efficient high-performance liquid chromatography/UV detection

The present work describes the validation of an easy, fast and efficient precolumn derivatization method for the quantification of oligosides, mannose and galactose obtained by degradation of galactomannans. This work combines an acid hydrolysis and an enzymatic degradation of natural galactomannans with the quantification of released residues by reversed-phase HPLC-UV, the most usual HPLC system in laboratories. In case of enzymatic degradation, mannotetraose has been detected and quantified for the first time, and an application to the evaluation of the galactosyl distribution in galactomannans is proposed. After an acidic hydrolysis, this method also allowed to obtain the mannose/galactose (M/G) ratio.     

High throughput screening of gene functions in mammalian cells using reversely transfected cell arrays: review and protocol

Reversely transfected cell microarrays (RTCM) have been introduced as a method for parallel high throughput analysis of gene functions in mammalian cells. Hundreds to thousands of different recombinant DNA or RNA molecules can be transfected into different cell clusters at the same time on a single glass slide with this method. This allows either the simultaneous overexpression or--by using the recently developed RNA interference (RNAi) techniques--knockdown of a huge number of target genes. A growing number of sophisticated detection systems have been established to determine quantitatively the effects of the transfected molecules on the cell phenotype. Several different cell types have been successfully used for this procedure. This review summarizes the presently available knowledge on this technique and provides a laboratory protocol.     

Theoretical study of specific hydrogen-bonding effects on the bridging P-OR bond strength of phosphate monoester dianions

It has been proposed that the driving force for the initial phosphoryl transfer step of protein tyrosine phosphatases (PTPases) could be activation of the substrate ROPO32- by means of an enforced hydrogen-bonding interaction between an aspartic general acid and the bridging oxygen atom O (Zhang et al. Biochemistry 1995, 34, 16088-16096). The potential catalytic effect of this type of interaction, with regard to P-OR bond cleavage, was investigated computationally through simple model systems in which an efficient intramolecular hydrogen bond can take place between a H-bond donor group and the bridging oxygen atom of the dianionic phosphate. The dielectric effect of the environment (epsilon = 1, 4, and 78) was also explored. The results indicate that this interaction causes significant lengthenings of the scissile P-OR bond in all media but with more extreme effects observed in the low dielectric fields epsilon = 1 and epsilon = 4. It is interesting that, in all cases examined, this interaction actually contributes to stabilize the reactant state while causing its P-OR bond to lengthen. Overall, our results support the idea that this specific hydrogen-bonding situation might well be used by PTPases as an important driving force for promoting phosphoryl transfer reactions through highly dissociative transition states.     

Hydrogen bonding and cation coordination effects in primary and secondary amines dissolved in carbon tetrachloride

Raman and infrared spectroscopy were used to investigate hydrogen-bonding interactions and cation coordination effects in solutions of lithium triflate (LiCF3SO3) dissolved in two primary amines, hexylamine (HEXA) and N,N-dimethylethylenediamine (DMEDA), and in a secondary amine, dipropylamine (DPA). Strong intermolecular hydrogen-bonding interactions and weaker intramolecular hydrogen-bonding interactions that occur only in DMEDA were spectroscopically distinguished in a comparison of pure HEXA, pure DMEDA, and the dilute solutions of these amines in CCl4. The spectroscopic shifts in intensity and frequency in the NH stretching region of DPA and DPA diluted in CCl4 were similar to those of HEXA. Dilute electrolyte solutions in carbon tetrachloride were prepared to analyze specifically the cation coordination effect. In these solutions, limited intermolecular hydrogen-bonding interactions are present, and the observed spectral shifts correspond primarily to the cation-induced shifts. The symmetric SO3 stretching region of the triflate anion was investigated to probe further the coordination of the cation. The local structures of the triflate ions and the amine groups in the electrolyte solutions dissolved in CCl4 are similar to the local structures in the corresponding amine-salt crystals previously reported by us.     

A fluorescent immunochromatographic test using immunoliposomes for detecting microcystins and nodularins

Microcystins (MCs), a group of cyclic heptapeptides produced by common cyanobacteria (blue green algae), cause both acute and chronic toxicity. Due to their toxicity, constant monitoring in drinking water, recreational waters as well as other potential exposure through ingestion of contaminated sea food, is very important. In this context, an immunochromatographic test (ICT) using a monoclonal antibody labeled with fluorescent liposomes (immunoliposomes) as tracer was developed, allowing a rapid and simple detection of a large number of MC and nodularin variants in field samples. The present ICT using immunoliposomes proved to be ten times more sensitive than the ICT using colloidal gold for labeling. To achieve quantitative measurement, this ICT was improved by including a stable signal on the control band allowing the expression of the results as a ratio of the fluorescence signals of the specific band versus the control band (SB/CB). Very low concentrations of MC-LR were detected in the analysis buffer (0.06 ng/ml), well below the guideline value of 1 ng/ml proposed by the World Health Organization (WHO), with a dynamic range from 0.06 to 1.5 ng/ml of MC-LR. This method was also validated using a hand-held commercial fluorometer (from ESE®), providing the same performances obtained via the analysis station (from Kodak®) used in our laboratory. Repeatability tests performed with both devices showed good accuracy (CV?Microcystis culture) was achieved using ICT, leading to similar results obtained via an EIA previously described. All these results demonstrate that this new fluorescent ICT could be used not only as a sensitive detection tool but also to quantify MCs in field samples.