Joop C. M. Waterval

Derivatization trends in capillary electrophoresis

This survey gives an overview of recent derivatization protocols, starting from 1996, in combination with capillary electrophoresis (CE). Derivatization is mainly used for enhancing the detection sensitivity of CE, especially in combination with laser‐induced fluorescence. Derivatization procedures are classified in tables in pre‐, on‐ and postcapillary arrangements and, more specifically, arranged into functional groups being derivatized. The amine and reducing ends of saccharides are reported most frequently, but examples are also given for derivatization of thiols, hydroxyl, carboxylic, and carbonyl groups, and inorganic ions. Other reasons for derivatization concern indirect chiral separations, enhancing electrospray characteristics, or incorporation of a suitable charge into the analytes. Special attention is paid to the increasing field of research using on‐line precapillary derivatization with CE and microdialysis for in vivo monitoring of neurotransmitter concentrations. The on‐capillary derivatization can be divided in several approaches, such as the at‐inlet, zone‐passing and throughout method. The postcapillary mode is represented by gap designs, and membrane reactors, but especially the combination of separation, derivatization and detection on a chip is a new emerging field of research. This review, which can be seen as a sequel to our earlier reported review covering the years 1991—1995, gives an impression of current derivatization applications and highlights new developments in this field.

Capillary electrophoresis as a versatile tool for the bioanalysis of drugs - a review

This review article presents an overview of current research on the use of capillary electrophoretic techniques for the analysis of drugs in biological matrices. The principles of capillary electrophoresis and its various separation and detection modes are briefly discussed. Sample pretreatment methods which have been used for clean-up and concentration are discussed. Finally, an extensive overview of bioanalytical applications is presented. The bioanalyses of more than 200 drugs have been summarised, including the applied sample pretreatment methods and the achieved detection limits.

Capillary electrophoretic bioanalysis of therapeutically active peptides with UV and mass spectrometric detection after on-capillary preconcentration

An earlier developed capillary electrophoresis (CE) system with an on‐capillary adsorptive phase is investigated for its suitability to quantitate low concentrations of angiotensin II and gonadorelin in plasma. An off‐line solid‐phase extraction is used for sample preparation. The on‐line preconcentration CE system allows multiple capillary volumes of sample solution to be injected, increasing the concentration sensitivity of CE with 3–4 orders of magnitude. Furthermore, possible influence of matrix salts can be ruled out by employing a rinsing step after sample application. Using short‐wavelength UV detection, reproducibility and linearity in the low nanomolar range were satisfactory. The capillary could be efficiently regenerated using a programmed between‐run rinsing procedure, allowing 20–30 large injections of sample extracts. Coating of the capillary improved the robustness of the method. Mass spectrometric detection via a previously reported sheathless interface increased the selectivity and sensitivity substantially. Recommendations are provided for the sample preparation process, the most critical part of the system. Further purification of the sample is required to allow the loading of larger sample volumes and to optimize the systems robustness.

Quantitative analysis of pharmaceutically active peptides using on-capillary analyte preconcentration transient isotachophoresis.

An on‐capillary adsorptive phase in combination with capillary electrophoresis (CE), frequently referred to as preconcentration CE, for quantitative analysis of low peptide concentrations was developed. The capillary containing the on‐line analyte preconcentrator can be constructed within 5 min from commercially available extraction disks. These disks contain poly(styrenedivinylbenzene) adsorbent particles incorporated in a matrix of inert Teflon, creating a mechanically stable sorbent. Therefore, no frits are needed in the capillary to hold the stationary phase in place. Several parameters, such as the required minimal elution volume, required elution strength, sample application speed or ionic strength, and the capacity were investigated and special interest was given to the quantitative properties of the method. Instead of nL injections, volumes up to a least 25 μL are possible, yielding improvements in detection limits of 3—4 orders of magnitude. The observed limit of detection for both model peptides was 20 pg, corresponding to a 20 μL injection of a 1 ng/mL solution of both model peptides. Using low‐wavelength UV detection, reproducibility and linearity in the low nanogram range were satisfactory. No influence of matrix salt concentrations was observed, extending the use of CE to all kinds of samples.

Robust and cost-effective capillary electrophoresis-mass spectrometry interfaces suitable for combination with on-line analyte preconcentration

This paper describes several successful cost‐effective attempts to couple capillary electrophoresis (CE) and mass spectrometry (MS) without make‐up flow or nebulizing gas. An in‐depth analysis of several interfaces using conductive spray tips was performed as well as an easy‐to‐prepare T‐junction with direct electrode contact, the latter being the most robust interface. No coating is necessary and the spray voltage is applied through a gold wire positioned at the gap between the separation and spray capillaries. The T‐junction interface is made by puncturing a small piece of transparent rubber. The on‐line preconcentration CE‐MS system allows immunoassay sensitivity, as is demonstrated by a calibration plot in the picomolar range for angiotensin II and gonadorelin. It also shows good reproducibility and has the ability of excellent automation. The secure electrical contact gives a constant spray quality, even with 100% aqueous separation buffers. The described setup has a wide applicability as is demonstrated by the analysis of larger peptides, such as insulin and cytochrome c. Detailed information is given on critical factors in the preparation of the described interfaces.

Comparison between transient isotachophoretic capillary zone electrophoresis and reversed-phase liquid chromatography for the determination of peptides in plasma

Low levels of peptide drugs in human plasma can be determined employing off‐line solid‐phase extraction, followed by capillary zone electrophoresis with UV detection. A bioanalytical procedure is presented, using gonadorelin and angiotensin II in human plasma as model compounds. The solid‐phase extraction method, based on a weak cation exchange mechanism, is able to remove interfering endogenous components from the plasma sample, extract the model peptides quantitatively, and give a possibility of concentrating the sample at the same time. Transient isotachophoretic conditions were kept to increase the sample loadability by about two orders of magnitude. Up to about 70% of the capillary was filled with the reconstituted extract, whereafter the peptides were selectively concentrated during the first 15 min. Subsequently, the concentrated sample zones were separated under capillary zone electrophoresis conditions, showing the techniques high resolution. For the model cationic peptides (gonadorelin, angiotensin II) good linearity and reproducibility was observed in the 20—100 ng/mL concentration range. A more extensive washing procedure permits quantitation of gonadorelin at the 5 ng/mL level. In comparison with a liquid chromatography analysis, superior mass sensitivity and separation are obtained with the transient isotachophoretic capillary zone electrophoresis method. Moreover, in this case equivalent sensitivity is achieved when it is directly compared with a liquid chromatography method with UV detection, keeping in mind that 60 times more sample is needed for the latter method. A further gain in sensitivity can be obtained when the analysis is combined with native fluorescence detection, as is demonstrated by combining liquid chromatography separation with fluorescence detection.