Challenges and applications of isotachophoresis coupled to mass spectrometry: A review.

Electrophoretic separations are of growing interest to tackle complex analytical challenges. Nevertheless, capillary electrophoresis, as the most common mode, still suffers from insufficient detection limits due to low capillary loadability. ITP is of growing interest as preconcentration method for capillary electrophoresis and is also interesting to be applied as an independent analytical method. While mass spectrometric detection is common for capillary electrophoresis, the combination of ITP with MS is still a niche technique. In this work, we want to give an overview on isotachophoretic effects in CE-MS and ITP-MS methods, as well as coupling techniques of ITP with CE-MS. The challenges and possibilities associated with mass spectrometric detection in ITP and its coupling to capillary electrophoresis are critically discussed.

Electromigrative separation techniques in forensic science: combining selectivity, sensitivity, and robustness.

In this review we introduce the advantages and limitations of electromigrative separation techniques in forensic toxicology. We thus present a summary of illustrative studies and our own experience in the field together with established methods from the German Federal Criminal Police Office rather than a complete survey. We focus on the analytical aspects of analytes' physicochemical characteristics (e.g. polarity, stereoisomers) and analytical challenges including matrix tolerance, separation from compounds present in large excess, sample volumes, and orthogonality. For these aspects we want to reveal the specific advantages over more traditional methods. Both detailed studies and profiling and screening studies are taken into account. Care was taken to nearly exclusively document well-validated methods outstanding for the analytical challenge discussed. Special attention was paid to aspects exclusive to electromigrative separation techniques, including the use of the mobility axis, the potential for on-site instrumentation, and the capillary format for immunoassays. The review concludes with an introductory guide to method development for different separation modes, presenting typical buffer systems as starting points for different analyte classes. The objective of this review is to provide an orientation for users in separation science considering using capillary electrophoresis in their laboratory in the future.

Label-free fluorescence detection of aromatic compounds in chip electrophoresis applying two-photon excitation and time-correlated single-photon counting.

In this study, we introduce time-resolved fluorescence detection with two-photon excitation at 532 nm for label-free analyte determination in microchip electrophoresis. In the developed method, information about analyte fluorescence lifetimes is collected by time-correlated single-photon counting, improving reliable peak assignment in electrophoretic separations. The determined limits of detection for serotonin, propranolol, and tryptophan were 51, 37, and 280 nM, respectively, using microfluidic chips made of fused silica. Applying two-photon excitation microchip separations and label-free detection could also be performed in borosilicate glass chips demonstrating the potential for label-free fluorescence detection in non-UV-transparent devices. Microchip electrophoresis with two-photon excited fluorescence detection was then applied for analyses of active compounds in plant extracts. Harmala alkaloids present in methanolic plant extracts from Peganum harmala could be separated within seconds and detected with on-the-fly determination of fluorescence lifetimes.

Nonaqueous capillary electrophoresis-mass spectrometry: a versatile, straightforward tool for the analysis of alkaloids from psychoactive plant extracts.

In this study we show that a nonaqueous capillary electrophoresis mass spectrometry (NACE-MS) method carefully optimized by a design of experiment can be applied to a very large number of alkaloids in different plant extracts. It is possible to characterize the pattern of the psychoactive alkaloids in several plant samples and preparations thereof, each presenting different challenges in their analysis. The method is shown to be able to separate structurally closely related substances, diastereomers and further isobaric compounds, to separate members of different alkaloid classes within one run and to tolerate significant matrix load. A comparison with methods presented in the literature reveals that a near-generic NACE-MS method for the fast profiling of alkaloids in forensically relevant plant samples has been developed.

Forensic analysis of mesembrine alkaloids in Sceletium tortuosum by nonaqueous capillary electrophoresis mass spectrometry.

The consumption of legal and illegal drugs follows an organic trend comparable to the current trend in food consumption. The investigation of such drugs is therefore of interest to characterize the active ingredients of plants and drug preparations. A new method of nonaqueous capillary electrophoresis coupled to mass spectrometry (NACE-MS) as a powerful tool for the separation of complex alkaloid mixtures in difficult matrices is presented in this study for the analysis of samples of Sceletium tortuosum and drug products called Kanna made thereof. The method was found to be suitable for the investigation of the alkaloid composition and relative quantification of the ingredients. It proved of value to separate a large number of isobaric compounds, most probably including diastereomers, double-bond isomers, and further structurally closely related compounds. A comparison of plant samples from different vendors, self-fermented samples, and products ready for consumption was made. The high separation power obtained allowed a better description of the chemotypic differences of plant samples as well as Kanna preparations compared to other methods presented in the literature so far. Thus, the use of the NACE-MS enables a new perspective on the alkaloid profile of Sceletium species.

Implementation of a design of experiments to study the influence of the background electrolyte on separation and detection in non-aqueous capillary electrophoresis-mass spectrometry.

Non-aqueous capillary electrophoresis (NACE) background electrolytes are most often composed of a mixture of methanol and acetonitrile (ACN) with soluble ammonium salts added as electrolyte. In this study on NACE-MS, we used a mixture of glacial acetic acid and ACN giving rise to an acidic background electrolyte (BGE) with a very low dielectric constant. Impressive changes in selectivity and resolution were observed for structurally closely related indole alkaloids including diastereomers upon addition of ammonium formate as electrolyte and upon variation of the solvent ratio. In order to obtain best separation and MS detection conditions and to reveal the influence of the parameters of the BGE on separation and detection and vice versa of the MS parameters on separation, an optimization strategy was employed using a design of experiments in a central composite design with response surface methodology. It was proven that at high electroosmotic flow conditions capillary electrophoretic separations and thus optimization can be realized without interference from the coupling to an MS system. Several significantly interacting parameters were revealed, which are not accessible with classical univariate optimization approaches. With this optimization, alkaloid mixtures from a plant extract of Mitragyna speciosa, containing a large number of diastereomeric compounds were successfully separated.