Abstract: A theophylline-binding polymer was prepared utilizing the technique of non-covalent molecular imprinting polymerization. The polymer obtained was packed in a high-performance liquid chromatography (HPLC) column, and the molecular recognition mechanism regulating the binding behavior was studied by frontal and zonal chromatography, determining the effects of pH and methanol on the affinity constant and concentration of the binding sites. The values of binding capacity found show the ability of the imprinted polymer to bind theophylline in aqueous buffer, even if with reduced affinity with respect to the values reported in literature for the same polymer in an organic medium. Hydrogen bonding was confirmed as leading interaction in the recognition mechanism. (C) 1997 Elsevier Science B.V