Dimethylsulfoxide oxidizes glutathione in vitro and in human erythrocytes: kinetic analysis by ¹H NMR

The interaction of dimethylsulfoxide (Me₂SO) with glutathione was investigated under non-equilibrium conditions in solution using ¹H NMR and in intact erythrocytes using ¹H spin-echo NMR. In solution the reaction was observed to follow second-order kinetics (Rate = k₁[glutathione][Me₂SO]) at 300 K pH 7.4, k_sol = 4.7 × 10^-5 mol ^-1 L¹ s^-1. In intact erythrocytes the rate constant for the cellular environment, k_cell, was found to be slightly larger at 8.1 × 10-5 mol-1 L¹ s^-1. Furthermore, the reaction of Me₂SO with erythrocyte glutathione showed a biphasic dependence on the Me₂SO concentration, with little oxidation of glutathione occurring until the Me₂SO concentration exceeded 0.5 mol L^-1. The results suggest that at lower concentrations, Me₂SO can be effectively removed, most probably by reaction with glutathione, which is regenerated by glutathione reductase, although preferential reaction with other cellular components (e.g., membrane or cellular thiols) cannot be ruled out. Thus the concentrations of Me₂SO that are commonly used in cryopreservation of mammalian cells (∼1.4 mol L^-1) can cause oxidation of intracellular glutathione.