Interaction of immunoglobulin G with modified chitosan

Chitosan beads were modified to include a spacer arm and end-capped with a caboxyethyl-group containing anionic ligand to generate a pseudo-bioaffinity support, further referred to as Ligosep Alpha® or LMCB. To better understand the force of interaction between the immunoglobulin G (IgG) and the chitosan-based pseudo-affinity support, the equilibrium dissociation constant (K_d) was determined by static binding isotherms, as a function of temperature, and by frontal analysis at different linear velocities. The maximum static binding capacity (Q_max) was found to be in the range of 60-70 mg IgG per mL of gel, and unaffected by temperature. The adsorption rate constant (k_a) was determined by a split-peak approach to be between 46 and 404 L mol^-1 sec^-1 depending on the linear velocity. A governing empirical equation relating k_a to the linear velocity (u) was deduced to be k_a = 44.4(u) + 9.3. The kinetic parameters of the chromatographic system indicated a good capacity but a low adsorption rate constant. LMCB was found to preferentially interact with the F_ab and the F_(ab)2 region of the HIgG molecule. This gives the further potential of adjusting the process for purifying specific paratopes under gentle chromatographic conditions.