Measurement of diffusion and partition coefficients of ferrocyanide in protein-immobilized membranes

True diffusion ( D m) and partition ( α) coefficients for the transport of potassium ferrocyanide through diaphorase (Dp)– and bovine serum albumin (BSA)–glutaraldehyde (GA) membranes with different cross-linking degrees of 1–8% GA concentrations immobilized on gold electrodes are investigated by using potential-step method and rotating-disk-electrode method. The thickness of dry and hydrated immobilized membranes is accurately measured by the focus-difference method with a reflection microscope. The thickness of hydrated Dp–GA and BSA–GA membranes are about 1.4 and 2.4 times that of dry membranes, respectively. In addition, the actual area of electrode surface is calculated by the charge amount of chemisorbed oxygen on gold electrode. Owing to the increase of swelling degree and net negative charge of the immobilized membranes, the values of D m and α for both of Dp–GA and BSA–GA membranes enlarge and decrease with increase of GA concentration, respectively. Furthermore, BSA–GA membranes possess greater D m and α than those of Dp–GA membranes due to the thinner thickness and the greater swelling degree of BSA–GA membranes.