Effects of Metal Ions, Alkylamines and Organic Solvents on the Separation of Biomolecules by Capillary Electrophoresis

The composition of the background electrolyte solution (BGE) employed in capillary zone electrophoresis (CZE) can strongly influence sample solubility and detection, native conformation of biopolymers, molecular aggregation, electrophoretic mobility and the electroosmotic flow (EOF). Consequently, selecting the proper composition of BGE is of paramount importance in optimizing the separation of the analytes in CZE. The appropriate selection of the buffer requires evaluating the physical-chemical properties of all components of the buffer system, including buffering capacity, conductivity, and compatibility with the detection system and with the sample. This communication discusses theoretical and practical aspects of the influence of the composition of the BGE on both the separation behaviour of biomolecules and the generation of EOF in bare fused-silica capillaries employed in CZE. Secondary equilibrium in solution between ionogenic analytes and components of the BGE can drastically affect the selectivity, which is based on differences in the electrophoretic mobility of the analytes, which depend on their effective charge-to-hydrodynamic radius ratios. This implies that selectivity is strongly influenced by the pH of the BGE as well as by the presence of suitable counterions and other interacting agents that may affect the net charge or/and the hydrodynamic radius of the analytes. The talk includes the discussion on the use of BGEs appositively tailored for separating proteins, peptides and other biomolecules by capillary electrophoresis with bare fused-silica capillaries. Most of the investigated solutions consist of buffering agents having the capability of controlling the protonic equilibrium in a wide pH range while preventing the undesirable interactions between basic analytes and the inner surface of bare fused-silica capillaries. Also discussed is the use of organic solvents and suitable additives incorporated into the BGEs traditionally employed in CZE.