Thermodynamic Characteristics of A Human Insulin-Deae-Dextran Complex Entrapped in Liposomes

Abstract

Binding parameters were measured for a study of in vitrocomplexation interaction of human insulin with diethylaminoethyl (DEAE) dextran polymer by a diaultrafiltration (rapid dialysis). The complex was found to have appearent affinity constants of 22.9, 3.2 and 1.5 M^-1 at 25°C, 37°C and 45°C respectively in 0.067 M phosphate buffer at pH 7.4. The complexation reaction was found to be exothermic with an enthalpy change (AH) of 26 kcalhole. Differential scanning calorimetry (DSC) was used to obtain the temperatures and enthalpies for the insulin denaturation process in the presence of the DEAE-dextran polymer and liposomal membranes. The human insulin-DEAE-dextran complex appears to have a higher denaturation temperature with a higher denaturation enthalpy than the free human insulin in 0.067 M phosphate buffer at pH 7.4 solution. The uncomplexed human insulin entrapped in dimyristoyl phosphatidylcholine (DMPC) liposomes and neutral liposomes (DMPC/cholesterol = 1 : 1) did not differ significantly in temperature and enthalpy of denaturation from the unentrapped protein. Conversely, the complexed insulin entrapped in liposomes indicated an unchanged denaturation temperature but a lower denaturation enthalpy compared to the complexed human insulin not entrapped. The complexed human insulin entrapped in positively charged liposomes prepared from DMPC/cholesterol/stearylamine in the molar ratio of 7:2:1 showed a slight change in denaturation temperatures and lower denaturation enthalpies than the complexed protein not entrapped whereas the uncomplexed human insulin gave three peaks of different denaturation temperatures and enthalpies.