The complex formation between Mg2+, Ca2+ and 1,2-dihydroxybenzene-3,5-disulphonate [Tiron: (H2L)(2-)] was investigated at 25.0 +/- 0.1 degrees C and in 0.1 mol.dm(-3) NaCl, KCl media. The potentiometric titrations were carried out, in the 1: 1 and higher mole ratios of M(II)-Tiron systems throughout the pH range 4.00 - 11.00. The stoichiometric protonation constants of Tiron were determined. The best reaction model comprises two complexes for each M(II)-Tiron system, of which one is monoprotonated (MHL)(+) type and the other (ML)(2-) type complex. The stoichiometric formation constants of the complexes (log beta) were equal for (MgHL)(+) and (CaHL)(+) complex species; log beta(2-)((MgL)) value is bigger than log beta(2-)((CaL)); nonetheless, Delta log beta values (the difference between log beta values) for (ML)(2-) type complexes of Mg2+ and Ca2+ in 0.1 mol.dm(-3) NaCl ionic medium is around (0.77 +/- 0.24), whereas it is around (0.33 +/- 0.18) in 0.1 mol.dm(-3) KCl ionic medium. Due to the higher covalency in Mg-OH2 bonds in [Mg(OH2)(6)](2+) type complex species than in Ca-OH2 bonds in [Ca(OH2)(n)](2+) complex species, Delta log beta value is not too big. In order to eliminate the most common error sources in biological media, the complex formations between Mg, Ca and Tiron should be taken into account. The effect of Tiron on Ca2+ is unrelated to its role as a radical scavenger. Therefore the effect of Tiron, a non-toxic chelator of a variety of metals, may need to be reconsidered as an antioxidant; since it binds Mg2+, Ca2+ and forms complex species. By means of the stoichiometric formation constants of Mg2+-Tiron and Ca2+-Tiron complex species, the equilibrium concentrations of free Mg2+, Ca2+ may be defined.