Influence of peak measurement parameters on the quality of chiral electrophoretic separations

The effect of nine peak measurement/analysis parameters on chiral capillary electrophoresis (CE), followed by UV detection, was studied. The parameters pertained UV-detection (detection wavelength, reference wavelength, and wavelength bandwidths), signal processing (data acquisition rate, type and amount of filtering) and peak detection (detection threshold and peak width). The influence of these factors on the chiral separation of dimethindene enantiomers was studied at two different concentrations (i.e., at high and low signal-to-noise (S/N) ratio) by the means of experimental designs. The electropherogram characteristics considered were the resolution between the two enantiomers, the peak areas, and the S/N ratio. A D-optimal design was first used as screening design to identify the most critical parameters. Afterwards, a modelling of the different responses as a function of these critical parameters was performed based on the results of a face-centered central composite design. The results showed that the signal-processing parameters should be carefully selected when developing a CE separation since very important variations in the separation, the S/N ratio and the peak area of the substances can occur by setting these parameters at different levels. The detection wavelength should also be carefully chosen for optimal peak area measurement. The role of these parameters becomes more important with decreasing concentration of the analytes (i.e., low S/N ratio). This study showed that the peak measurement/analysis parameters should be optimized as the chemical and physical parameters of a method. They also should always be well specified in order to allow a good transfer of a method from one instrument to another.     

Acylation des β-énaminoesters: Regio et stéréoformation des β-énaminoesters N- ou C-acylés

The reaction of acyl chlorides with cyclic five-membered β-enaminoesters gave exclusively N-acylated products while reaction of acyl chlorides with cyclic seven-membered β-enaminoesters gave only C-acylated products. In the case of cyclic six-membered β-enaminoesters, the reaction of acyl chlorides gave a mixture of N-acylated and C-acylated products.     

Synthesis of 1,1,1 - trifluoro-2-penten-4-one

The 1,1,1-trifluoro-2-penten-4-one has been prepared from trifluoroacetaldehyde by two different ways. Its condensation with a cyclohexanone enamine leads, by annelation, to 4-trifluoromethyl-Δ-1,9 (and Δ-9,10)-octalin- 2-one.     

Influence of polyelectrolyte coating conditions on capillary coating stability and separation efficiency in capillary electrophoresis

Polyelectrolytes are widely used in capillary electrophoresis as coating agents of silica capillaries to prevent adsorption phenomena and improve the repeatability of peptide and protein analysis. A systematic study of the coating experimental conditions has been carried out to optimize coating stability and performance. The main experimental parameters studied were the type and concentration of polyelectrolytes used in several monolayer and multilayer coatings, the ionic strength of coating and stabilizing solutions, and the procedures used for coating and capillary storage. Electroosmotic flow magnitude, direction and repeatability were used to monitor coating stability. Coating ability to limit adsorption was investigated by monitoring variations of migration times, time-corrected peak areas and separation efficiency of test peptides. Capillary-to-capillary and batch-to-batch reproducibility was also studied. In addition, the separation performance of polyelectrolyte coatings were compared to those obtained with bare silica capillaries.     

LIF detection of peptides and proteins in CE

CE- and microchip-based separations coupled with LIF are powerful tools for the separation, detection and determination of biomolecules. CE with certain configurations has the potential to detect a small number of molecules or even a single molecule, thanks to the high spatial coherence of the laser source which permits the excitation of very small sample volumes with high efficiency. This review article discusses the use of LIF detection for the analysis of peptides and proteins in CE. The most common laser sources, basic instrumentation, derivatization modes and set-ups are briefly presented and special attention is paid to the different fluorogenic agents used for pre-, on- and postcapillary derivatization of the functional groups of these compounds. A table summarizing major applications of these derivatization reactions to the analysis of peptides and proteins in CE-LIF and a bibliography with 184 references are provided which covers papers published to the end of 2005.     

Precision study on capillary electrophoresis methods for metacycline

A CE method for metacycline (MTC) determination was investigated in an inter-laboratory experiment. Many problems were encountered in this study, most of which were related to the transfer of the method to different CE equipment. The reported problems could be classified into different categories: problems related to the precision, to the parameters in the protocol, and to the MTC peak shape. As the peak shape problem was partially responsible for the poor precision, a new CE method was developed in order to obtain a good MTC peak shape on all equipment. The precision of this new method for MTC determination was examined in an intermediate precision study, where the influence of the factors "time" and "equipment" was investigated. Although the new method could be transferred to different instruments, the precision remained poor mainly due to the contributions of the between-replicate and the between-injection variances.     

Use of coated capillaries for the electrophoretic separation of stereoisomers of a growth hormone secretagogue

The diastereoisomeric separation of peptidomimetics of hexarelin, a strong growth hormone secretagogue, in CE has been studied. Highly sulfated‐γ‐CD was found to be an appropriate selector for the separation of the stereoisomers. However, non‐repeatable analyses were obtained on bare fused silica capillary due to the progressive adsorption of the analytes on the capillary wall. Two types of polyelectrolyte coating agents were tested to prevent this phenomenon. Coating with neutral polyethylene oxide was found to be efficient but resulted in a very long analysis time (about 40 min). Coating with cationic poly(diallyldimethylammonium) chloride was found both to prevent analyte adsorption, reduce analysis time and alter separation selectivity. EOF measurement revealed that the highly sulfated‐γ‐CDs were strongly adsorbed on the poly(diallyldimethylammonium) chloride coating surface yielding a stable strong cathodic EOF, which considerably reduced analysis time (about 12 min). Very good repeatability of analysis was obtained (RSD_{migration time}<1%).     

Isothiocyanato boron dipyrromethenes--the first BODIPY analogues of fluorescein isothiocyanate

Two boron complexes of 5-phenyldipyrromethenes bearing isothiocyanate groups on the phenyl ring have been synthesized for the first time. The utility of these new fluorescence probes for labeling biologically relevant proteins is demonstrated on two monoclonal antibodies that bind to antigens overexpressed on cancer cells. Spectral comparison of the two structures reveals significant photophysical differences, including bathochromically shifted excitation and emission bands, increased molar absorptivity and a large increase in fluorescence quantum yield of approximately 10 times. Differences in photophysical parameters are linked to hindered rotation of the phenyl ring in one of the probes.