Diffusional kinetics of monoclonal antibody, transferrin and bovine serum albumin on Sephacryl S-200 HR gel

Abstract

The adsorption and diffusional kinetics of three macromolecules, namely bovine serum albumin (BSA), transferrin and monoclonal antibody (McAb), on Sephacryl S-200 high resolution gel were studied by the pulse-response chromatographic technique in a column of diameter 1.6 cm and length 8.5 cm. Experimental results showed that the voidage ε and axial mixing (2D L /V) in the packed column were 0.432 and 0.05 cm based on the response to a pulse input of blue dextran 2 000 000 and D 2O solution respectively. The equilibrium constant K and pore diffusivity D p were 0.23 and (0.60 – 1.02) × 10 −7 cm 2 s −1 for BSA (molecular weight, 67 000) 0.17 and (0.62 – 1.0) × 10 −7 cm 2 s −1 for transferrin (molecular weight, 95 000) and 0.095 and (0.53 – 0.85) × 10 −7 cm 2 s −1 for McAb (molecular weight, 150 000 IgG). A van't Hoff plot of ln K vs. 1/T gave a straight line with zero slope and hence zero heat of sorption for all three components. This implies that no adsorption occurs between the molecules and the packing, and that the partition of solute in the bulk solution and pore solution is entirely controlled by diffusional forces.