Protolysis and Mg2+ binding of methylthymol blue

Spectrophotometric pH titrations of Methylthymol Blue (MTB) with tetramethylammonium hydroxide between pH 3.05 and 13.45 in the presence of a pH buffer, and cation titrations with MgCl2 (and to some extent CaCl2) between pH 6.08 and 8.49 have been carried out to determine apparent macroscopic stability constants of protolysis and cation coordination. The spectral properties provide evidence that MTB binds up to two cations in 1:1 and 1:2 stoichiometry. Based on a model considering protolysis of MTB and its complex formation with Mg2+, log K values are obtained of three states of the free ligand (LH(3)(3-), LH(2)(4-), LH(5-)) being 6.9, 11.0 and 13.7, of two states of its 1:1 Mg2+ complex (LMgH(2)(2-), LMgH(3-)) being 6.7 and 10.4, as well as of one state of its 1:2 complex (LMg(2)H(-)) being 6.3. Regarding Mg2+ binding, three different states of the 1:1 complex (LMgH(2)(2-), LMgH(3-), LMg(4-)) with log K values of 2.8, 7.1 and 10.4 and two states of the corresponding 1:2 complex (LMg(2)H(-), LMg(2)(2-)) with log K values of 1.5 and 5.6 are determined. The data refer in general to an actual or reduced ionic strength of 0.1 M and 25 degrees C. The quantitative determination of the binding parameters has been performed employing the described formalism, especially suitable for the present experimental conditions.