Zone formation ion-pair reversed-phase liquid chromatography: IV. Optimization of peak retention in step-gradient elution with introduction of competing ions

Step-gradient elution by introduction of a con-ion into an ion-pair reversed-phase system was studied. The analytes were organic acids and amines and were used as counter ions in eluents also containing phosphate buffer (pH 4 or 6)—methanol (1:1). The magnitude of the retention decrease obtained for the analytes was dependent on the hydrophobicity and concentratin of the co-ion and on a temporary decrease in the counter-ion concentration on introduction of the co-ion. When some prerequisites were fulfilled, a simple equation could be used to predict the retention volume, and then the separation time could be optimized. Extremely narrow peaks were obtained. The peak width was dependent on the co-ion concentration in the second eluent and also on the efficiency of the column, which is responsible for the bandbroadening of the breakthrough front of the co-ion.