From the assumptions of this binding process and Langmuir’s binding theory, the interaction between 5-fluorouracil and human serum albumin (HSA) has been investigated by the nano-watt-scale isothermal titration calorimetry and the circular dichroism (CD) spectrometry at 298.15 K. The results show that there are two classes of binding sites on the protein HSA for the 5-fluorouracil. For the first class of binding, the binding site number is N＝71±0.1, the binding constant is K＝(1.46±0.016)×10⁵ L•mol^－1, the binding enthalpy is ΔH＝(39.61±0.220) kJ•mol^－1, the binding entropy is ΔS＝(231.68±0.025) J• mol^－1•K^－1, and the binding Gibbs free energy is ΔG＝(－29.48±0.030) kJ•mol^－1. This binding is an entropy driven process, and the hydrophobic interaction is the main motive-force for the process. For the second class of binding, the binding site number is N＝140±0.2, the binding constant is K＝(1.49±0.032)×10⁵ L•mol^－1, the binding enthalpy is ΔH＝(－19.31±0.103) kJ•mol^－1, the binding entropy is ΔS＝(34.30±0.055) J•mol^－1•K^－1, and the binding Gibbs free energy is ΔG＝(－29.53±0.041) kJ•mol^－1. This binding is an enthalpy-entropy synergically driven process, and the hydrogen bond and electrostatic interactions are the main motive-force for the process. The analytical results of circular dichroism (CD) spectra show that the interactions between 5-fluorouracil and HSA changed the relative contents of secondary structure units of protein HSA in these two classes of binding processes. The thermodynamic effects of the binding system are integrated results which come from different interactions in the binding process. The conformations of the protein HSA underwent changes induced by the anti-tumor drug 5-fluorouracil in the solution medium of this binding system.

Key words: isothermal titration calorimetry, circular dichroism spectrometry, 5-fluorouracil, human serum albumin