[Studies on retention behaviour of biopolymers on silica-based binding stationary phase in the mobile phase of acetic acid-water].
The reversed-phase chromatographic method consisting of acetic acid-water as a new mobile phase for separating biopolymers on ODS column is presented. The new mobile phase may overcome the weak points of irreversible adsorption in chromatographic separation to some degree and difficulty in concentrating proteins with lyophylization and reduce the three-component (organic solvent, ion-pairing agent and water) of typical mobile phase to two component system. Based on the retention and Z (the number of solvent molecules involving in the displacement from stationary-phase surface), log I (a constant relating to the affinity of 1 mol of solute to the bonded stationary phase), and j (a constant relating to the affinity of 1 mol of solvent to the bonded stationary phase), the elution strength of acetic acid-water mobile phase was found to be stronger than that of methanol-water-TFA. From a good linear relationship between log I and Z, it can be concluded that the interaction force between proteins and RPLC stationary phase with acetic acid-water as mobile phase is non-selective hydrophobic interaction forces. In addition, the molecular conformations of protein in acetic acid-water and methanol-trifluoroacetic acid-water were found to be different.