Capillary-channeled polymer fibers as the stationary phase for high-performance liquid chromatography separations.

摘要

High-performance liquid chromatography (HPLC) has become one of the most widely used separation techniques in use today. Applications include the quality control of raw materials, intermediates, and finished products as well as the separation, isolation, and purification of proteins. The work described in the first part of this dissertation involves the fundamental studies of the hydrodynamic and separation characteristics of capillary-channel polymer (C-CP) fibers as the stationary phase in HPLC columns. The second part of this dissertation involves applications of C-CP fiber stationary phases for the separations of proteins in ion-exchange (IEC) and hydrophobic interaction (HIC) modes.; Initially, a study of the hydrodynamic (A-term) aspects of polypropylene and PET (poly(ethylene terephthalate)) C-CP fibers, ranging in diameter of ∼35 to 65 mum, was undertaken to illustrate the effects of packing density and column inner diameter on peak width and asymmetry. The elution of uracil at linear velocities ranging from 5 to 40 mm/s was used to determine the optimum linear velocity in terms of "apparent" plate height. Because uracil is unretained, issues regarding mass transfer and polymer/solute are negligible and thus only the A-term is of significance. The van Deemter A-term was evaluated as a function of fiber packing density (≈ 0.3 g/cm3 to 0.75 g/cm3) for columns of 4.6 mm inner diameter (i.d.) and at constant packing densities for 1.5 mm, 3.2 mm, 4.6 mm, and 7.7 mm i.d. columns. The results showed that C-CP fiber packing density, as well as column inner diameter have a significant effect on the peak asymmetry and flow homogeneity.; The use of nylon C-CP fiber as the stationary phase in mixed mode ion-exchange chromatography (IEC)/reversed-phase columns enabled the separation of a three-protein mixture. In IEC, binding is due to the interaction of a charged solute to a stationary phase of opposite charge to achieve electroneutrality. Elution is achieved by the employment of a gradient from low to high salt concentrations, thus increasing the concentration of competing ions in the mobile phase. (Abstract shortened by UMI.)