Optimisation and validation of an automated solid phase extraction technique coupled on-line to enzyme-based biosensor detection for the determination of phenolic compounds in surface water samples

Summary A fully integrated screening system for phenolic compounds was developed incorporating on-line solid phase extraction, fractionation and biosensor detection. Two different types of biosensors, solid graphite and carbon paste electrodes incorporating the enzyme tyrosinase, were compared and used in the screening system. Interfacing of the solid phase extraction and fractionation with the biosensor detection was given special attention since the biosensors were not compatible with the organic modifier used for desorption of phenols from the solid phase extraction step. The system was validated with conventional analytical techniques. Surface water samples from the Ebro river were spiked with 1,10, and 25g L^–1 of catechol, phenol,p-cresol, respectively. Three out of seven samples were spiked and the correct samples were identified, containing phenols equivalent to the spiked concentrations.     

Nouveaux heterocycles tricycliques synthese de derives de l'aza-2 pyrrolo[1,2-a]indole

The synthesis of aza-2 pyrrolo[1,2-a]indole derivatives was achieved by a 1,3-dipolar cycloaddition reaction of dimethylacetylenedicarboxylate with a mesoionic ion.     

Comparative bioanalytical study of3H-deramciclane in dog plasma, using a gas chromatography-nitrogen-selective detection (GC-NPD), a new GC-radiochemical detection (GC-RD) and a liquid scintillation method

Summary A new, highly sensitive and selective gas chromatography method, using radiochemical detection (GC-DR) was developed for the selective determination of³H-labelled deramciclane and its N-desmethyl metabolite in dog plasma. Inter-day accuracy and precision, as well as system suitability of the GC-RD method was investigated during the method validation. The calibration curve was proved to be linear (r=0.9986) in a wide concentration range (13–1000 ngeqv mL⁻¹) The lower limit of quantitation (LLOQ) was 13.7 ngeqv mL⁻¹, and the limit of the detection (LOD) was 1 ngeqv mL⁻¹. Using this new GC-RD method, plasma levels of³H-labelled deramciclane and its metabolite were determined in dogs, after the administration of a single 10 mg kg⁻¹ oral dose. Pharmacokinetic curves and the calculated pharmacokinetic parameters were compared to those obtained using a previously elaborated gas chromatography-nitrogen selective detection method (GC-NPD) and to those obtained by measuring the plasma level of total radioactivity (liquid scintillation counting, LSC). Pharmacokinetic curves and the calculated pharmacokinetic parameters obtained with the two different gas chromatography detection methods (NPD and RD) showed good correlation. Comparison of these results to those acquired by total radioactivity measurement demonstrated that deramciclane was intensively metabolised. Moreover, the biological half-life (t ₁ ^2/β ) of the unknown metabolites proved to be more than a magnitude longer than the half-life of the parent compound or that of N-desmethyl metabolite. Presented at: Balaton Symposium on High-Performance Separation Methods, Siófok, Hungary, September 3–5, 1997.     

Effect of brain lesions on [3H]ohmefentanyl binding site densities in the rat striatum and substantia nigra.

We have recently demonstrated that [3H]ohmefentanyl, a non-peptidergic opioid ligand which was suggested to cross the blood brain barrier in contrast to other peptidergic opioid ligands, bound not only to mu opioid receptor sites but also to sigma sites. In order to examine whether [3H]ohmefentanyl can be used as a marker for mu sites, we investigated the effects of brain lesions on [3H]ohmefentanyl binding site densities, as compared with [3H][D-Ala2, MePhe4, Gly-ol5]enkephalin ([3H]DAGO), a selective mu ligand. These binding site densities were measured by quantitative autoradiography in the rat striatum and substantia nigra, two brain structures known to contain a high density of mu receptors, following lesions of the nigro-striatal dopaminergic pathway and striatal intrinsic neurons. Following unilateral nigral lesion with 6-hydroxydopamine, [3H]ohmefentanyl binding site densities were decreased in the patches (-35%) and matrix (-20%) of the ipsilateral striatum and in the lesioned substantia nigra pars compacta (-49%). Unilateral striatal lesion with quinolinic acid induced 72%, 61% and 50% decreases in [3H]ohmefentanyl binding in the patches and matrix of the lesioned striatum and in the ipsilateral substantia nigra pars reticulata, respectively. Similar results were obtained in the binding of [3H]DAGO. Indeed, a significant linear correlation was observed between [3H]ohmefentanyl and [3H]DAGO binding site densities. Therefore, mu opioid receptors may be mainly located on intrinsic neurons in the striatum, dopaminergic cell bodies in the substantia nigra pars compacta and nerve terminals of striatal efferents in the substantia nigra pars reticulata.(ABSTRACT TRUNCATED AT 250 WORDS)     

Synthesis of branched oxime-linked peptide mimetics of the MUC1 containing a universal T-helper epitope

Our goal was to develop mimics of MUC1, highly immunogenic to induce an efficient immune response against the tumor-associated form of MUC1, and sufficiently different from the natural antigen to bypass the tolerance barrier in humans. With the aim of obtaining a well-defined peptide construct as a means of evoking the precise immune responses required in immunotherapy, we synthesized artificial mimics of the MUC1 protein composed of two MUC1 repeat units of inverse orientation and a universal T-helper epitope. To synthesize these heteromeric peptide constructs, we followed a convergent approach using chemoselective ligation based on oxime chemistry. A stem peptide was first synthesized bearing two orthogonally masked aldehydes. After successive deprotection, two oxime bonds can be specifically generated. The proposed strategy proved to be concise and robust, and allowed the synthesis of the tri-branched protein in a very satisfactory yield. The different constructs were tested for their ability to generate antibodies able to recognize the MUC1 protein.     

Synthesis and Structure of a 1,3-alternate-Thiacalix[4]arene diamide Derivative

A novel receptor possessing two complexation sites and bearing 1,3-alternate conformation based on thiacalix[4]arene, confirmed by single crystal X-ray analysis, was prepared. The tetrathiacalix[4]arene diamide shows strong intramolecular hydrogen bonding. The binding behavior towards K⁺ and halides has been examined by ¹H NMR titration experiments.     

The molecular geometries of some cyclic nitramines in the gas phase

The structural parameters of 1,3,5-trinitro-1,3,5-triazacyclohexane (RDX), (CH₂NNO₂)₃, 1,3-dinitro-1,3-diazacyclopentane (DDCP), CH₂(CH₂NNO₂)₂, andN-nitropyrrolidine (NP), (CH₂)₄NNO₂, have been determined by electron diffraction.The six-membered ring of RDX has a chair form with axial positions of the nitro groups and close to planar bond geometry of the amine nitrogen atoms. The overallC ₃ symmetry of the molecule is in agreement with the experimental data.The conformation of the five-membered ring in DDCP is a half-chair ofC ₂ symmetry, while that in NP is an envelope ofC _S symmetry. The nitro groups are in equatorial positions in both molecules. The conformations of pyrrolidine and imidazolidine cycles show interesting features.The pyramidal geometry of the amine nitrogen atom bonds flattens in going from pyrrolidine andN-chloropyrrolidine to NP and DDCP and then to RDX and to dimethylnitramine (DMNA), (CH₃)₂NNO₂.     

Application of pressurized sample preparation methods for the analysis of steels and copper alloys

Pressurized sample preparation devices (High Pressure Asher, Pressurized Microwave Digestion system, compared with a PTFE decomposition vessel) were used to dissolve certified metal alloy samples (steel, copper) for ICP analysis. Based on the results of the analysis it was established that both up-to-date devices can be advantageously applied to quickly and quantitatively dissolve metal alloy samples. To dissolve the samples, two different kinds of acid mixtures (A: nitric and hydrochloric acid; B: nitric and hydrochloric and sulphuric and phosphoric acid) were used. The sample preparation is simpler and less time-consuming than the earlier commonly used methods, sample loss and degree of contamination are also reduced. Steel samples containing tungsten, titanium and niobium (less than 0.5%) can only be analyzed using a mixture of the four acids. By dissolving steel samples in the nitric and hydrochloric acid mixture, the concentration of their most common elements (Cr, Ni, Mn, V, Cu) as well as their S and P content can be determined. Copper alloy samples can be dissolved quickly by the pressurized microwave decomposition device using hydrochloric acid and diluted (1:1) nitric acid.     

The characteristics of integrins expression in decidualized human endometrial stromal cell induced by 8-Br-cAMP in in vitro

Integrins are heterodimeric glycoproteins that have been found to undergo dynamic temporal and spatial changes in the endometrium during the menstrual cycle and in early pregnancy. Specificity of integrins is known to be different in human endometrial stromal cells and decidual cells. These shifts of integrins suggested to play an important role in embryo implantation and can be modulated by progesterone, cAMP derivatives, and cytokines. The mechanisms of decidualization and its precise physiological role are still not clearly understood and in vitro systems could provide an alternative that overcomes limitations of studying such complex biological phenomena in vivo at the time of implantation. This study was undertaken to establish an in vitro model system for human decidualization using 8-bromo-cAMP and to investigate the characteristics of stromal integrin expression in vitro by 8-Br-cAMP. Endometrial stromal cells were isolated and cultured, and then were induced to decidualize by 0.5 mM 8-Br-cAMP for 15 days. Immunofluorescence staining and flow cytometric analyses of the integrin subunits (alpha1, alpha4, alpha5, alpha6, beta1 and alphavbeta3) were performed at day 9. In the presence of 8-Br-cAMP, the staining intensity of alphavbeta3 was significantly higher than control and measurements for alpha1, alpha4, alpha5, alpha6, and beta1 were similar. Immunofluorescent localization of the integrins reflected the differences obtained from the flow cytometric analyses described above. In summary, the expression of alphavbeta3 integrin increased in stromal cells in vitro decidualized by 8-Br-cAMP and this up-regulation of alphavbeta3 integrin expression during decidualization might influence on human implantation.