Mass transfer mechanisms in Hyper D media for chromatographic protein separation

Abstract

In order to elucidate the mass transfer mechanisms that allow improved performance for the separation of proteins of the chromatographic packing material Q Hyper D (BioSepra, Villeneuve la Garenne, France), when compared with traditional packing materials, several experiments were carried out using a commercial HPLC column, with BSA and myoglobin as test proteins. First, elution chromatographic runs under unretained conditions at several flowrates and protein concentrations were made. The HETP vs. superficial velocity plot is almost a plateau, except for the low flowrate region. Elution experiments were also carried out under weakly retained conditions (with salt concentration 0.3 M NaCl) in the linear region of the adsorption equilibrium isotherm. Using Rodrigues' equation, the experimental initial slope and plateau of a Van Deemter plot enable the determination of the effective diffusivity and particle permeability respectively. Then, breakthrough experiments under retaining conditions were run at several feed concentrations and flowrates. The experimental adsorption equilibrium isotherm is rectangular; using a simplified fixed-bed adsorber model for rectangular isotherms, an apparent (augmented) diffusivity is calculated as a function of flowrate. The determined augmented diffusivities are shown to follow the dependence on velocity established by Rodrigues et al.: this clearly shows that intraparticle convection is present as a mass transfer mechanism, since the real intraparticle diffusivity is independent of flowrate.