Displacement countercurrent chromatography

Abstract

The present invention provides a new method of pH-zone-refining countercurrent chromatography that can be operated in a manner analogous to displacement chromatography. The method uses a retainer base (acid) in the stationary phase to retain analytes in the column and a displacer acid (base) to elute the analytes in the decreasing (or increasing) order of pK.sub.a and hydrophobicity. The elution produces a train of highly concentrated rectangular solute peaks with minimum overlap. To use pH-zone refining CCC in a displacement mode, the mobile and stationary phases are switched. Thus, the original eluent becomes a retainer to retain analytes in the stationary phase, and the original retainer acid becomes a displacer to displace the analytes from the stationary phase to the mobile phase at the back of the solute bands. The present method provides a distinct advantage over the pH-zone-refining countercurrent chromatography in the normal mode in that the eluted compound is provided as a salt-free acid or base in an organic solvent which can easily be evaporated. Additionally, the displacement mode of pH-zone-refining countercurrent chromatography is amenable to a ligand-affinity separation which may cover a broader range of analytes including nonionizable compounds. As with the normal mode, the present method can be used on a preparative scale.