Electrophoresis of solutes in aqueous two-phase systems.

Abstract

Electrophoresis of solutes was studied in aqueous two-phase systems, concentrating on the special behavior in the interfacial region. Moving boundary electrophoresis was examined in a free fluid U-tube apparatus. Zone electrophoresis was investigated in two-phase systems which were gelled by the addition of acrylamide, which was subsequently polymerized. The size and nature (concentration or dilution) of polarizations which were found to occur was found to depend on the magnitude of the equilibrium partition coefficient of the solute in the two-phase system, as well as the direction of migration across the interface. These polarizations are in addition to those commonly known to occur near regions where electrophoretic flux changes radically, such as near interfaces. They can be a direct result of the requirement for equilibrium across the interface, as demonstrated by our experiments. Models were constructed to numerically simulate this behavior, which accounted for unsteady state electrophoresis and diffusion of multiple proteins or other amphoteric solutes. Two cases were explored, one requiring instantaneous solute equilibration across the interface, the other allowing for a resistance to mass transfer here. All models demonstrated a characteristic noted in experimental studies, concentration at interfaces when electrophoresis is from equilibrium preferred phase towards non-preferred phase. Furthermore, the equilibrium model correctly predicted the complex relationship between partition coefficient, direction of migration, and moving boundary or zone electrophoresis, which causes differences in the polarizations observed in these various systems. The simulation could also quantitatively estimate the width of the polarized region to within an order of magnitude, in comparison with experimental results, while hampered by a lack of mobility data for solutes in solutions containing polymers.