CAPILLARY ZONE ELECTROPHORESIS IN A HYDRODYNAMICALLY CLOSED SEPARATION SYSTEM WITH ENHANCED SAMPLE LOADABILITY

Some phenomena linked with capillary zone electrophoresis (CZE) perfor med in a hydrodynamically closed separation system with enhanced sampl e load capacity via the use of capillary tubes of larger I.D.s were st udied. Calculations of the plate heights for varying amounts of the an alytes loaded onto 50 and 300 mu m I.D. columns under identical CZE se parating conditions revealed that the analytical advantages of using c olumns of larger I.D, include significantly reduced contributions of e lectromigration dispersion. At higher concentrations of the carrier el ectrolytes this gain, however, can be partially lost due to increased thermal dispersive effects. Calculated resolutions for a varying ratio of a pair of the analytes loaded onto 50 and 300 mu m I.D. columns fa voured the use of the latter I.D. in situations when the ratio of the analytes was higher than ca. 10(2):1. CZE experiments were carried out in a 300 mu m I.D. capillary tube made of fluorinated ethylene-propyl ene copolymer (FEP) with a porous cellophane membrane serving as a hyd rodynamic barrier to prevent a flow of the solution in the separation compartment due to a pressure difference between the electrode vessels . Movement of the membrane was found to be a source of undesired flows in the separation compartment, which adversely affected both the repr oducibilities of migration times of the analytes and their separation efficiencies. Mechanical support eliminated these problems. Dispersive phenomena associated with electroosmosis in the closed separation com partment were effectively suppressed by using high molecular weight de rivatives of water soluble polymers (methylhydroxyethylcellulose and p olyethyleneglycol) in the carrier electrolyte solutions. Examples from the separations of various groups of analytes (synthetic food coloura nts, some inorganic anions, alkali and alkaline earth metal cations an d glycoforms of erythropoietin) are given to illustrate a practical ut ility of the studied CZE approach.