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Silica-based monolithic capillary columns modified to zwitterionic stationary phase for hydrophilic interaction liquid chromatography
Moravcová, Dana ; Planeta, Josef ; Kahle, Vladislav ; Horká, Marie ; Roth, Michal
Zwitterionic monolithic capillary columns intended for isocratic gradient hydrophilic interaction chromatography (HILIC) separations are introduced. Silica-based capillary columns (150 mm x 0.1 mm) were prepared by acidic hydrolysis of tetramethoxysilane in the presence of polyethylene glycol and urea. The modification by a 3-(trimethoxysilyl)propyl methacrylate and then by a zwitterionic [2-(methacryloyloxy)ethyl]-dimethyl-(3-sulfopropyl)-ammonium hydroxide to HILIC stationary phase bearing sulfoalkylbetaine groups on its surface followed. Prepared columns were characterized in HILIC separation mode employing mobile phase containing 10% (v/v) of 5 mM ammonium acetate pH = 4.5 in acetonitrile. Comparison with the commercially available ZIC-HILIC® column (Merck SeQuant®) under the same separation conditions using a mixture of aromatic carboxylic acids as a sample was done on the basis of separation efficiency of tested columns as well as retention factors and peak asymmetry of individual solutes.
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Using of divergent flow isoelectric focusing and capillary liquid
Duša, Filip ; Moravcová, Dana ; Kahle, Vladislav ; Šlais, Karel
Separation of analytes from complex matrices play important role in their further identification. In this article we propose two-dimensional separation method of bovine serum albumin tryptic peptides. At the first dimension peptides are separated by divergent flow isoelectric focusing followed by reverse-phase liquid chromatography as the second dimension. Due to composition of input mixture it is possible to use autofocusing mode in first dimension. In this mode peptides are focused without addition of carrier ampholytes. Chromatograms from the second dimension were processed into contour map. Acquired data showed good separation efficiency of isoelectric focusing. Autofocusing mode enabled better resolution of peptides with close isoelectric points.
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