The acid–base and complex-formation equilibria of [Ru(H₂dtpa)(H₂O)], where dtpa = diethylenetriaminepentaacetate, with a series of bio-relevant ligands having various functional groups, viz. thiol, amine, imidazole and carboxylate, were investigated using potentiometric and spectrophotometric techniques. The pK_a values for [Ru(H₂dtpa)(H₂O)] were found to be 2.28 and 3.48 for the uncoordinated carboxylic acid groups and 8.83 for the coordinated water molecule. The complexes formed are of 1:1 stoichiometry and their formation-constants were determined. The effect of dioxane on the acid–base and complex-formation equilibria of the Ru^III complex was studied. The displacement reaction of coordinated NO by the investigated ligands showed that thiols can compete with NO in their reaction with [Ru^III(dtpa)(H₂O)]²⁻. The results reveal that the Ru^III complex is deactivated as a NO scavenger by thiolate ligands.