The interaction of 6-p-toluidinylnaphthalene-2-sulfonate (TNS) with beta-cyclodextrin was investigated in 0.1 M phosphate buffer at pH 7.4 by fluorescence spectroscopy. Using the fluorescence enhancement of TNS in the presence of beta-cyclodextrin, the thermodynamic parameters for the formation of two kinds of the inclusion complex (molar ratio of beta-cyclodextrin to TNS = 1:1 and 2:1) were determined as follows: delta G degree 1 (1:1 complex) = -20.0 kJ mol-1 at 25 degrees C, delta H degree 1 = -19.6 kJ mol-1, delta S degree 1 = -1.7 J mol-1 K-1, delta G degree 2 (2:1 complex) at 25 degrees C = -6.14 kJ mol-1, delta H degree 2 = -2.80 kJ mol-1, delta S degree 2 = 16.7 J mol-1 K-1. From the thermodynamic parameters, the main driving force for the 1:1 inclusion complex formation was considered to be the van der Waals-London dispersion force, while the contribution of the hydrophobic interaction was small. Also, the hydrogen bonding was suggested to contribute to the inclusion complex formation. The main driving force for the 2:1 inclusion complex formation was the hydrophobic interaction. Also, from the measurements of proton nuclear magnetic resonance spectra and studies with Corey-Pauling-Koltun atomic models, the probable structure was determined and discussed in connection with the thermodynamic parameters.