New mu-oxo-diferric complexes have been designed for hydrolysis of phosphodiesters. To mimic the diiron active site of purple acid phosphatase, a combinatorial method has been used to select complexes containing two distinct iron coordination spheres. The introduction of a bidentate ligand, a substituted phenanthroline (L) into complex 1, [Fe2O(bipy)(4)(OH2)(2)](NO3)(4), generates in solution the complex [Fe2O(bipy)(3)(L)(OH2)(2)](NO3)(4) as shown by ESI/MS and H-1 NMR studies. The latter complex was found to be 20-fold more active than complex 1. On the basis of kinetic studies, we demonstrated that the complex [Fe2O(bipy)(3)(L)(OH)(OH2)](NO3)(3) was the active species and the reaction proceeded through the formation of a ternary complex in which one iron binds a hydroxide and the second, the substrate. At nonsaturating concentrations of the substrate, the increased activity with increased methyl substituents in L was due to an increased affinity of the complex for the substrate. The activity of [Fe2O(bipy)(3)(33'44'Me-2-Phen)(OH2)(2)](NO3)(4) [33'44'Me(2)Phen = 3,3',4,4'-dimethyl-1,10-phenanthroline] was found to be comparable to that reported for Co(III) or Ce(IV) complexes.