This paper deals with the separation of isomers of nitroaniline by liquid chromatography using the ligand-exchange technique. The chromatographic separations were performed on the ligand-exchanger sporopollenin. The sporopollenin used as support of stationary phase was modified with carboxylated-ethylenediamine matrix and was loaded with cobalt(II) ions. Using the column packed with cobalt(II) loaded carboxylated diaminoethyl sporopollenin [Co(II)-CDA-E-S], the retention behavior of 3- and 4-nitroanilines was investigated. The mobile phase used, was a mixture of 0.05 M NH4OH in ethanol-water. The resolution was strongly affected by the presence of ammonium hydroxide in the mobile phase and a concentration of 0.05 M was shown to be necessary for the separation of analytes. To study the effects of temperature on the resolution. column runs were also performed at various temperatures (15-60 degrees C). With increasing temperature. a decreased interaction between the solutes and the ligand-exchanger was observed. Consequently. the best results were obtained using a mixture of 0.05M NH4OH in ethanol-water (10:90. v/v) as the mobile phase at a column temperature of 35 degrees C. Ligand-exchange chromatography on the Co(II)-CDAE-S could be a useful alternative method for the separation of nitroaniline. (c) 2005 Elsevier B.V. All rights reserved.