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Pure Appl. Chem., 2001, Vol. 73, No. 4, pp. 745-759


Measurement and analysis of results obtained on biological substances with differential scanning calorimetry (IUPAC Technical Report)

Hans-Jürgen Hinz1 and Frederick P. Schwarz2

1 Institut für Physikalische Chemie, Westfalische Wilhelms-Universität, Schlossplatz 4/7 D-48149, Münster, Germany
2 Center for Advanced Research in Biotechnology/National Institute of Standards and Technology, 9600 Gudelsky Drive, Rockville, Maryland 20850, USA

Abstract: Differential scanning calorimeters (DSCs) have been widely used to determine the thermodynamics of phase transitions and conformational changes in biological systems including proteins, nucleic acid sequences, and lipid assemblies. DSCs monitor the temperature difference between two vessels, one containing the biological solution and the other containing a reference solution, as a function of temperature at a given scan rate. Recommendations for DSC measurement procedures, calibration procedures, and procedures for testing the performance of the DSC are described. Analysis of the measurements should include a correction for the time response of the instrument and conversion of the power vs. time curve to a heat capacity vs. temperature plot. Thermodynamic transition models should only be applied to the analysis of the heat capacity curves if the model-derived transition temperatures and enthalpies are independent of the DSC scan rate. Otherwise, kinetic models should be applied to the analysis of the data. Application of thermodynamic transition models involving two states, two states and dissociation, and three states to the heat capacity vs. temperature data are described. To check the operating performance with standard DSCs, samples of 1 to 10 mg mL­1 solutions of hen egg white lysozyme in 0.1 M HCl-glycine buffer at pH = 2.4 ± 0.1 were sent to six different DSC laboratories worldwide. The values obtained from proper measurements and application of a two-state transition model yielded an average unfolding transition temperature for lysozyme of 331.2 K with values ranging from 329.4 to 331.9 K, and an average transition enthalpy of 405 kJ mol­1 with values ranging from 377 to 439 kJ mol­1. It is recommended that the reporting of DSC results be specific with regard to the composition of the solution, the operating conditions and calibrations of the DSC, determination of base lines that may be model-dependent, and the model used in the analysis of the data.