Principles and Practice of Chromatography - Chromatography Applications > Liquid Chromatography Applications > Ionic Interaction Chromatography > Page 85
The column was 25 cm long, 4.6 mm in diameter, and packed with silica gel particle (particle diameter 5 mm) which gave a maximum efficiency at the optimum velocity of 25,000 theoretical plates. The mobile phase consisted of 76% v/v n-hexane and 24% v/v 2-propyl alcohol at a flow-rate of 1.0 ml/min.. The steroid hormones are mostly weakly polar, consequently, the separation on silica gel, will be based primarily on polarity. The silica, however, was heavily deactivated by a relatively high concentration of the moderator 2-propyl alcohol and thus the silica gel surface would be covered with isopropanol and thus the interacting surface would be virtually pure 2-propyl alcohol. Whether the interaction is by sorption or displacement is difficult to determine. It is likely that the early peaks interacted by sorption and the late peaks possibly by displacement.
Ionic Interaction Chromatography
Ionic interaction chromatography, or ion chromatography as it is usually called, is typically carried out employing ion exchange resins as the stationary phase. There are some silica based ion exchange materials available, but the bonded silicas tend to be unstable in the presence of high salt concentrations and at extremes of pH. As a consequence, they have very limited areas of application. Alternatively, the polystyrene divinyl benzene cross-linked polymers, are extremely stable to a wide range of salt concentrations and can function well within the pH range of 2.0 to 12.0. An obvious application area for ion exchange chromatography is in the separation of all types of anions and cations. Metal cations and inorganic anions are all separated predominantly by ionic interactions with an ion exchange resin. Organic acids and bases, however, would be retained by mixed interactions, as dispersive and polar interactions will take place between the solute molecules and the aromatic nuclei and the aliphatic side chains of the polymer base. The separation of simple acids and bases require that the mobile phase is buffered appropriately according to the pKa of the salts so that dissociation occurs and the ions are free to interact with the stationary phase. By employing mobile phase additives and using novel operating conditions, ionic interactions can be used to separate a far wider range of materials than simple organic and inorganic anions and cations.