The acid dissociation constants (pK(a)) of 7-hydroxyflavone, 3-hydroxyflavone and 5-hydroxyflavone, which are practically insoluble compounds in water but of great biological and physicochemical interest, were determined by UV-visible spectroscopy in ethanol-water and acetonitrile-water solutions, varying the solvent relative permittivity in the interval of 61 to 75, at constant ionic strength (0.050 mol.kg(-1)) and temperature (298.15 K). The pK(a) values of the compounds increase as the permittivity of the reaction medium decreases. Correlations were established between the pK(a) values and empirical parameters of the solvents, such as the relative permittivity and hydrogen-bond donor capacity. These equations allow the interpretation of the solvent effect on the acid-base equilibria and the determination of pK(a), values at 298.15 K in pure water. The ordering of the pK(a) values was 7-hydroxyflavone (7.28) < 3-hydroxyflavone (8.68) < 5-hydroxyflavone (11.75). In addition, the dissociation constants were also calculated by means of DFT methods (B3LYP/6-311+G(2d,p) level of theory), employing several thermodynamic cycles. The solvent effect on the optimized structures in the gas-phase was evaluated using the polarizable continuum model. A good agreement was observed between the theoretical and experimental pK(a), values. Finally, the experimental ordering in the acidity of the hydroxyflavones was explained using natural bond orbital analysis.