Abstract
Adsorption kinetics of bovine serum albumin (BSA) and bovine hemoglobin (bHb) to Cibacron Blue 3GA (CB) modified Sepharose CL-6B has been studied. The effects of liquid-phase ionic strength and CB coupling density on the uptake rates of these two proteins in Tris–HCl buffer (pH 7.5) were evaluated by effective pore diffusivity derived from a pore diffusion model. The results showed that despite their similar molecular masses and sizes, the effects of aqueous-phase ionic strength and CB density on the effective pore diffusivities of BSA and bHb were distinctly different. The effective pore diffusivity of BSA to CB-Sepharose increased significantly with decreasing CB density and increasing liquid-phase ionic strength. This was considered due to the decrease in electrostatic repulsion between the BSA and CB molecules of like charge. That is, the increase in ionic strength and the decrease in CB coupling density reduced the electrostatic hindrance effect on BSA diffusion to CB-Sepharose, facilitating the hindered pore diffusion. In contrast, because of the higher isoelectric point of bHb (7.0) compared to BSA (4.7), bHb suffered little electrostatic hindrance effect during its diffusion to CB-Sepharose. Therefore, the effective pore diffusivity of bHb was unchanged with the change in liquid-phase ionic strength and CB coupling density.
Published Version
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