CHARACTERIZATION OF DIFFERENT RP-HPLC COLUMNS BY A GRADIENT ELUTION TECHNIQUE

The literature contains no unified testing procedure for evaluation an d characterization of reversed-phase (RP) columns of different pore si zes (surface area) and containing different types of ligand. In our la boratory a testing procedure has been developed using gradient elution under standardized conditions independent of column dimensions and ap plicable to both narrow-pore and wide-pore columns. Six wide-pore and four narrow-pore columns were investigated and compared. The test solu tes were selected to cover a wide range of chemical properties. The ev aluation and characterization of the columns was performed on the basi s of k(app) retention factors. Principal components analysis (PCA) was performed to reveal similarities and differences among the columns an d test substances. The factors obtained characterize the columns accor ding: to the extent of various interactions such as hydrophobicity, HB (hydrogen bond)-donor acidity and HB-acceptor basicity.