The lactate-dependent enhancement of hydroxyl radical generation by the Fenton reaction

The effect of lactic acid (lactate) on Fenton based hydroxyl radical ((OH)- O-.) production was studied by spin trapping, ESR, and fluorescence methods using DMPO and coumarin-3-carboxylic acid (3-CCA) as the (OH)-O-. traps re spectively. The (OH)-O-. adduct formation was inhibited by lactate up to 0. 4 mM (lactate/iron stoichiometry = 2) in both experiments, but markedly enh anced with increasing concentrations of lactate above this critical concent ration. When the H2O2 dependence was examined, the DMPO-OH signal was incre ased linearly with H2O2 concentration up to 1 mM and then saturated in the absence of lactate. In the presence of lactate, however, the DMPO-OH signal was increased further with higher H2O2 concentration than 1 mM, and the sa turation level was also increased dependent on lactate concentration. Spect roscopic studies revealed that lactate forms a stable colored complex with Fe3+ at lactate/Fe3+ stoichiometry of 2, and the complex formation was stri ctly related to the DMPO-OH formation. The complex formation did not promot e the H2O2 mediated Fe3+ reduction. When the Fe3+-lactate (1:2) complex was reacted with H2O2, the initial rate of hydroxylated 3-CCA formation was li nearly increased with H2O2 concentrations. All the data obtained in the pre sent experiments suggested that the Fe3+-lactate (1:2) complex formed in th e Fenton reaction system reacts directly with H2O2 to produce additional (O H)-O-. in the Fenton reaction by other mechanisms than lactate or lactate/F e3+ mediated promotion of Fe3+/Fe2+ redox cycling.