Examination of kinetic effects in the high-performance liquid affinity chromatography of glycoproteins by stopped-flow and pulsed elution methods

Using a concanavalin A high-performance liquid affinity chromatography system, the effect of slow solute desorption from the silica-bound ligand on protein recovery and peak shape has been investigated. The desorption of glycoproteins from the immobilized concanavalin A was found to be many times slower than the desorption of monovalent carbohydrates. To overcome the kinetic limitations of this chromatographic system, a novel method of protein elution, stopped-flow elution, was developed and used to ovalbumin and to examine the kinetic "sluggishness" of the desorption process. The stopped-flow elution method allows the quantitative recovery of purified glycoprotein in a volume of approximately 0.5 ml.