Kinetics of the cleavage of two phosphate diesters, bis(4-nitrophenyl) phosphate and 2-hydroxypropyl 4-nitrophenyl phosphate and a triester, 4-nitrophenyl diphenyl phosphate, in the presence of Mg(ii), Ca(ii) and Sr(ii) were studied in 90% vol. DMSO at 37 °C. The alkaline hydrolysis of the triester was inhibited by all cations, but with both phosphodiesters strong catalytic effects were observed. Potentiometric titrations of metal perchlorates by Bu4N(OH) revealed formation of M2(OH)3+, M(OH)+, M(OH)2and M2(OH)5−species. Rate constants for phosphodiester cleavage by individual species were obtained from analysis of rate–concentration profiles. Observed first-order rate constants in the presence of 1–2 mM Mg(ii) or Ca(ii) in neutral and weakly basic solutions were 108–1011times higher than those for background hydrolysis at the same pH while in water additions of up to 50 mM metal produced <100-fold accelerations. Possible structures of DMSO solvated catalyst–substrate complexes were modeled by DFT calculations with Mg(ii). The increased catalytic activity in 90% DMSO is attributed to stronger association of hydroxide ions and anionic phosphodiesters with metal ions and to preferable solvation of cations by DMSO, which creates favorable for reaction anhydrous microenvironment in the coordination sphere of the catalyst. [ABSTRACT FROM AUTHOR]