Critique of methods for estimating heats in isothermal titration calorimetry

Isothermal titration calorimetry data recorded on a MicroCal/Malvern VP-ITC instrument for water-water blanks and for dilution of aqueous solutions of BaCl2 and Ba(NO3)(2) are analyzed using Origin software, the freeware NITPIC program, and in-house algorithms, to compare precisions for estimating the heat per injection q. The data cover temperatures 6-47 degrees C, injection volumes 4-40 mu L, and average heats 0-200 mu cal. For water-water blanks, where baseline noise limits precision, NITPIC and the in-house algorithm achieve precisions of 0.05 mu cal, which is better than Origin by a factor of 4. The precision differences decrease with increasing vertical bar q vertical bar, becoming insignificant for vertical bar q vertical bar > 200 mu cal. In its default mode, NITPIC underestimates vertical bar q vertical bar for peaks with incomplete return to baseline, but the shortfall can be largely corrected by overriding the default injection time parameter. The variance estimates from 26 dilution experiments are used to assess the data variance function. The results determine the conditions under which weighted least squares should be used to estimate thermodynamic parameters from ITC data.