The application of affinity capillary electrophoresis (ACE) to the study of molecular interactions is reviewed. ACE appears to be a sensitive, versatile and convenient tool to obtain reliable data on binding constants and stoichiometries of interacting systems using the Hummel-Dreyer method and variants thereof. A powerful feature is the possibility to analyze simultaneously the affinity of a large number of compounds for the same ligand, making it a promising tool for the screening of large combinatorial libraries.

The literature contains no unified testing procedure for evaluation and characterization of reversed-phase (RP) columns of different pore sizes (surface area) and containing different types of ligand. In our laboratory a testing procedure has been developed using gradient elution under standardized conditions independent of column dimensions and applicable to both narrow-pore and wide-pore columns. Six wide-pore and four narrowpore columns were investigated and compared. The test solutes were selected to cover a wide range of chemical properties. The evaluation and characterization of the columns was performed on the basis of kapp retention factors. Principal components analysis (PCA) was performed to reveal similarities and differences among the columns and test substances. The factors obtained characterize the columns according to the extent of various interactions such as hydrophobicity, HB (hydrogen bond)-donor acidity and HB-acceptor basicity.

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