Nanoparticle-based capillary electrochromatography
Sammanfattning: This thesis concerns a highly efficient nanoparticle-based capillary electrochromatography separation technique and its coupling with mass spectrometric detection. From chromatographic theory, it is well known that the separation efficiency increases with decreasing particle size. However, as the particle size decreases, the pressure required to push a liquid through the column also increases. This increase in back pressure results in problems both in packing and use of the columns. Therefore, there is a limitation on how small the particles used in traditional chromatography can be. In this thesis, a method is described that is based on capillary electrochromatography using a suspension of nanoparticles as a pseudostationary phase. With these nanoparticle suspensions, separation efficiencies as high as 1.1 million plates per meter were obtained. The technique can be used with very small particles, and there is rather an upper limit on particle sizes due to loss of suspension stability. The nanoparticles have a charge-to-size ratio that is different from that of the analytes. Consequently, the velocity of the nanoparticles is different from the velocity of the analytes under an applied electric field. The analytes are thus separated due to the difference in their partitioning between the liquid phase and the nanoparticle phase. In the thesis, three fundamentally different nanoparticle types have been synthesised and used. Molecularly imprinted polymer nanoparticles have been used for enantiomer separations, highly charged nanoparticles have been used for ion chromatographic separations, and hydrophobic nanoparticles have been used for reversed-phase separations. Thus, the technique can be applied to a broad range of different separation problems, and some examples are demonstrated here. The presented technique has several advantages over traditional chromatography performed with immobilised interaction phases. The nanoparticle suspension is exchanged after every separation, and thus, problems due to adsorption of sample components to the interaction phase are absent. Also, inexpensive bare fused silica capillaries are used and clogging or other damage to the capillary does not result in high costs. Therefore, the technique has great potential for analysis of complex samples such as blood plasma and urine. Determination of theophylline in spiked blood plasma was performed. Also, changes in interaction phase, in terms of type and amount, can be made without changing the column or capillary. Combinations of different nanoparticles can be used to resolve complex samples in a novel multimode fashion. This technique was used for simultaneous separation of the enantiomers of two different analytes. Compared to other separation techniques that employ relatively low molecular weight pseudostationary phases, such as proteins, surfactants and cyclodextrins, the nanoparticle-based interaction phases was shown to have improved compatibility with mass spectrometric detection. With an orthogonal electrospray ionisation interface, the analytes could be separated from the nanoparticles prior to mass spectrometric detection. Sensitive detection of the analytes without degradation of analyte signal intensity was achieved.
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