A new method is evaluated whereby enantioselective binding is examined by use of steady-state fluorescence anisotropy measurements. A theoretical treatment is presented that relates fluorescence anisotropy measurements to chiral selectivity and allows the estimation of thermodynamic parameters of chiral recognition. It is shown that the natural logarithm of the ratio of anisotropy values of two enantiomers varies as a function of temperature in a manner where the slope and intercept are directly related to the differential enthalpy (Delta Delta H degrees) and entropy (T Delta Delta S degrees) of the enantioselective interaction. The developed method was evaluated by examining the enantioselective interactions of several chiral molecules with beta-cyclodextrin and a molecular micelle.