The first volume profiles for complex formation of alpha-cyclodextrins (alpha-CD) with diphenyl azo dyes (S) are presented as a new approach in understanding inclusion phenomena. The following dyes were selected: sodium 4-(4-diethylaminophenylazo)benzenesulfonate (1), sodium 4-(3-carboxy-4-hydroxy-5-methylphenylazo)benzenesulfonate (2), sodium 4-(4-hydroxy-3,5-dimethylphenylazo)benzenesulfonate (3), and sodium 2-hydroxy-3-methyl-5-(4-sulfamoylphenylazo)benzoate. The behavior of the dyes alone were first studied in aqueous solutions to rule out any competition reaction. Under the experimental conditions used for the stopped-flow kinetic studies, it has been proved that only monomeric species are present (no aggregation of the dye is formed by pi-pi stacking interactions). NMR experiments and kinetic evidences have shown that only directional binding of the dye via the sulfonate/sulfonamide group through the wide rim of the alpha-cyclodextrin was possible. The 1:1 complex was the only stoichiometric species formed. The inclusion reactions for the four selected dyes were characterized by a two-step kinetics described by a first fast step that yields the intermediate, S.alpha-CD*, followed by a slower rearrangement to form the final complex, S.alpha-CD. 2D NMR experiments served for a molecular dynamics calculation leading to a structural representation of the intermediate and final complexes. An interpretation of the volume profiles obtained from high-pressure stopped-flow kinetic experiments have not only confirmed the so far proposed mechanisms based on "classical" kinetic investigations but offered a new focus on the inclusion process. The inclusion mechanism can be summarized now as follows: the complexation begins with an encounter of the dye and alpha-cyclodextrin mainly due to hydrophobic interactions followed by a partial desolvation of the entering head of the dye. The latter interacts with the two "activated" inner water molecules of the free host and their complete release is delayed by the primary hydroxy group barrier of the alpha-CD. At this first transition state, a squeezed arrangement develops inside the cavity inducing a negative activation volume (Delta V-1,(f)double dagger approximate to -8 to -24 cm(3) mol(-1)). The subsequent intermediate is characterized by a total release of the two inner water molecules and interactions of the dye head with the primary hydroxy groups of the host in a trapped-like structure (Delta V(1)degrees approximate to -11 to -4 cm(3) mol(-1)). The latter interactions and concurrent tail interactions with the secondary hydroxy groups of the host lend at different extents to a strained conformation of the host in the second transition state (Delta V-2,(f)double dagger approximate to -2 to -16 cm(3) mol(-1)). In the final complex, the head of the dye is totally rehydrated as it protrudes from the primary end of the host cavity which can now adopt a released conformation (Delta V(2)degrees approximate to +3 to +6 cm(3) mol(-1) vs +17 cm(3) mol(-1) for 1).