D.G. Khachidze, Y.L. Kalandadze and J.R. Monaselidze

It is shown that the denaturation process of a medicinal preparation of insulin in dilute solution covers the temperature range 50–96 ºC. The excess heat capacity (D C_p) increases linearly with temperature and is equal to 0.29 J g ^–1 K^–1 at 62 ºC. A significant peak with T_d = 81.5 ºC, D H_d = 43.4 J/g and D T_d = 13.5 ºC is observed in the temperature range 62–96 ºC. Calculation of the ratio D H_d ^cal / D H_d ^eff of calorimetric to effective (van’t Hoff) enthalpies gives the value 0.83±0.1 which is fairly close to 1.0, indicating that the denaturation process of insulin proceeds as usually observed for compact globular proteins. The melting process of a medicinal preparation of Zn(II)-insulin, which is not an optically transparent solution, has three stages with T_d¹ = 77.0, T_d² = 82.5 and T_d³ = 91.5 ºC; all these stages correspond to various aggregation states of the protein. Hence it is inferred that Zn(II)-insulin exists in three different energetic states depending on concentration: in the concentration range 0.1–0.5%, D H_d decreases from 25.3 to 16.0 J/g; in the concentration range 1.5–40% is characterized by a constant value of D H_d equal to 16.0 J/g; in the concentration range 40–80%, D H_d drops monotonously from 16.0 to 5.2 J/g. It is evinced that this decrease of D H_d is connected with disruption of the protein hydration shell.

Keywords: calorimeter, denaturation, enthalpy, insulin