The dissociation constants of representative loop (furosemide), thiazide (chlorthiazide and trichlormethiazide) and potassium sparing (amiloride) diuretics in 10, 30, 40, 50 and 70% (w/w) acetonitrile-water mixtures at 298.15 K were determined, in accordance with IUPAC procedures. The variation in pK a values over the whole composition range can be explained by preferential solvation and the structural features in acetonitrile-water mixtures. Correlations between pK a values and various bulk and solvatochromic properties of the solvents were calculated. The linear solvation energy relationship (LSER) method was applied. The resulting equations allowed us to calculate pK a values for the diuretics studied in any acetonitrile-water mixture up to 70% (w/w) and should help to clarify the acid-base behaviour of diuretics in the widely-used acetonitrile-water mixed solvents.