Energetics of Ca2+-EDTA interactions:: calorimetric study

The interaction between Ca2+ and EDTA has been studied using isothermal titration calorimetry to elucidate the detailed mechanism of complex formation and to relate the apparent thermodynamic parameters of calcium binding to the intrinsic effects of ionization. It has been shown that Ca2+ binding to EDTA is an exothermic process in the temperature range 5-48 degrees C and is highly dependent on the buffer in which the reaction occurs. Calorimetric measurements along with pH-titration of EDTA under different solvent conditions shows that the apparent enthalpy effect of the binding is predominantly from the protonation of buffer. Subtraction of the ionization effect of buffer from the total enthalpy shows that the enthalpy of binding Ca2+ to EDTA is - 0.56 kcal mol(-1) at pH 7.5. The Delta H value strongly depends on solvent conditions as a result of the degree of ionization of the two amino groups in the EDTA molecule, but depends little on temperature, indicating that the heat capacity increment for metal binding is close to zero. At physiological pH values where the amino groups of EDTA with pK(a) = 6.16 and pK(a) = 10.26 are differently ionized, the coordination of the Ca2+ ion into the complex leads to release of one proton due to deprotonation of the amino group having pK(a) = 10.26. Increasing the pH up to 11.2, where little or no ionization occurs, leads to elimination of the enthalpy component: due to ionization,while its decrease to pH 2 leads to its increase, due to protonation of the two amino groups. The heat effect of Ca2+/EDTA interactions, excluding the deprotonation enthalpy of the amino groups, i.e. that associated with the intrinsic enthalpy of binding, is higher in value (Delta(b)H(o) = - 5.4 kcal mol(-1)) than the apparent enthalpy of binding. Thus, the large Delta G value for Ca2+ binding to EDTA arises not only from favorable entropic but also enthalpic changes, depending on the ionization state of the amino groups involved in coordination of the calcium. This explains the great variability in Delta(H) obtained in previous studies. The ionization enthalpy is always unfavorable, and therefore dramatically decreases Ca2+ affinity by reduction of the enthalpy term of the stability function. The origin of the enthalpy and entropy terms in the stability of the Ca2+-EDTA complex is discussed. (C) 1999 Elsevier Science B.V. All rights reserved.