Gert Desmet

Use of kinetic plots for the optimization of the separation time in ultra-high-pressure LC

The kinetic-plot approach, in which experimental t(0) and N-values are extrapolated to the performance at maximum system pressure by increasing the column length, was validated by coupling 250×3 mm columns packed with 3 μm particles. The extra-column volume introduced by coupling columns could be neglected with respect to the peak volumes. Plate numbers of up to 132,000 were experimentally achieved by coupling four columns. The maximum deviation between the experimental and predicted plate numbers was 7% for two coupled columns, and decreasing to 0.1% for four coupled columns. Kinetic plots were used to find the conditions to separate a critical pair, with a preset value for the effective plate number, in the shortest possible time. For high-efficiency separations yielding 100,000 effective plates, the optimum critical-pair retention factor was around 4.5. Kinetic plots are presented to find the optimal column length to obtain the fastest possible 100,000 effective-plate separation, taking into account the effect of mobile-phase viscosity on column pressure, and consequently the optimum column length.

Design and fabrication of a biomedical Lab-on-Chip system for SNP detection in DNA

A Lab-on-Chip system is proposed, capable of SNP (Single Nucleotide Polymorphism) detection in DNA. One of the core components is an advanced filter consisting of an ordered array of Si micro-pillars enabling fast and effective separation of 5 DNA segments with different length using chromatographic techniques. Also a dedicated micro-pump is fabricated based on conductive polymer actuation, generating the required high pressure to sustain the fluid flow through the total system. For the detector, a known detector principle is applied, but pronounced miniaturization is carried out in order to make a small and portable system.

Design and implementation of injector/distributor structures for microfabricated non-porous pillar columns for capillary electrochromatography

A previously proposed foil definition is applied in the design of injector/distributor structures for solid microfabricated column structures for capillary electrochromatography. In addition to a typical bifurcated distributor, an optimized design alternative with two different configurations is experimentally evaluated. Optimized designs yielded a flat profile for the injected sample with a maximum of 3% variation from the mean width, while it went up to 18% for the typical bifurcated distributor. The implemented electrokinetic injection approach enabled controlling the volume of the injected sample accurately without sacrificing the compactness of the device design. The width of the injected sample was directly proportional to the injection time, namely 165 and 218 μm base widths were obtained for 0.6 and 0.8s of feeding, respectively. Reducing the external porosity of the distributor by 85% compared to the typical design, optimized distributors caused a decrease in the mean flow velocity of up to 70%. However, having a flat initial plug shape enabled the separation of a mixture of Coumarin 440, 460, 480 and 540 at 1 mm downstream of the injection point in 80s, while it was even not possible to detect the C440 signal for a typical bifurcated design.

Fluorescence and absorbance measurements for chromatographic analysis using a miniaturized micro-optical detection unit

We present a micro-optical detection unit for both laser induced fluorescence and absorbance analysis in fused silica capillaries, which can be used for chromatographic applications. The detection system is designed by means of non-sequential ray tracing simulations and prototyped by means of Deep Proton Writing. Such a prototyped master component is afterwards replicated by means of elastomeric moulding and vacuum casting. In a proof-of-concept demonstration the prototyped master micro-optical unit is used for the detection of various concentrations of coumarin dyes. The detection limit (SNR=3.3) achieved measures 0.6nM for fluorescence analysis and 12μM for absorbance measurements in capillaries with an inner diameter of 150μm. We discuss the optimization of different measurement parameters of the detectors in the setup in order to achieve accurate, fast and sensitive measurements.

Tolerance analysis of a micro-optical detection unit for fluorescence and absorbance measurements in lab-on-a-chip micro-channels for chromatographic applications

The integration of optical components on microfluidic devices is needed for downscaling analytical processes to portable, integrated and low-cost lab-on-a-chip systems for point-of-care applications. We have developed a micro-optical detection unit for both laser induced fluorescence and absorbance analysis in fused silica capillaries for microfluidic chromatographic applications. We present the use of non-sequential ray tracing simulations to design the system and to perform a tolerance analysis to define theoretically for each parameter in the system the acceptable fabrication and alignment errors. The system is prototyped using Deep Proton Writing and characterized by means of an optical non-contact profiler, in order to check for every parameter if the realized alignment and fabrication errors do not exceed the theoretically acceptable tolerance ranges. These measurements show that Deep Proton Writing is appropriate for the fabrication of the designed micro-optical detection system. In addition the tolerance study shows for which parameters the alignment is most critical. Finally we demonstrate by means of optical simulations that the same micro-optical design can be applied in different materials (index of refraction between 1.3 and 1.5) and used for sensing fluorescence of a variety of molecules in a wide spectral window (from 400nm up to 1550nm).

Chapter 1:General Overview of Fast and High-resolution Approaches in Liquid Chromatography

Over the past 50 years, column packings have evolved from irregularly shaped silica particles with sizes of 30–100 µm to spherical particles with diameters of 3–5 µm and even less than 2 µm.1 The tendency to develop smaller particle sizes has in essence been driven by the urge to obtain more efficie...

A novel optical detection system for chromatography applications

In this work we describe the design of a confocal microscope based optical system which can be used for two types of miniaturized fluid-chromatographic experiments: first of all for the study of shear-driven fluid separations in microchannels and secondly for the study of peak-broadening phenomena in pressure-driven fluids in the same type of microchannels. For the first type of measurements fluorescing dyes (coumarins) are used while for the latter type photo-active molecules (uncaged dyes) like for example fluorescein molecules are needed. We designed the detection system in such a way that by making some small changes to the system we can swap from one application to the other. After calculating the specifications of the optical components, we simulated the system with the sequential ray-tracing-software Solstis and built the system. Finally we did some preliminary experiments which demonstrated the working principle of our setup.