Protective effect of alpha-keto acids on the oxidative hemolysis

We studied antioxidative properties of various concentrations (0.3-2 mM) of biochemically important alpha-keto acids: pyruvate, alpha-ketoglutarate, oxaloacetate, glyoxylic acid as aldehyde acid and also 2-methyl-thiazolidyne-2,4-dicarboxylic acid and their effect on the oxidative hemolysis of human erythrocytes induced by hydrogen peroxide (H2O2). Normal erythrocytes proved to be very resistant to oxidative damages, so the high concentration of H2O2 (10 mM) as well as the presence of natrium azide, a catalase inhibitor, was necessary. The levels of malonyldialdehyde (MDA), reactive oxygen species (ROS) and hemoglobin (Hb) released were evaluated as the measure of red cell peroxidative hemolysis. Pyruvate, at the lowest used concentration (0.3 mM), caused the inhibition of lipid peroxidation (MDA) and a drop in the level of ROS, as well as a diminution of the degree of hemolysis and the effects were stronger than those of other alpha-keto acids. At the highest (2 mM) concentration, the protective effect against oxidative damage of all the investigated alpha-keto acids was similar and amounted to nearly 50% in relation to the control sample. On the contrary, in the case of aldehyde acid, e.g. glyoxylic acid, no protective effect in the same range of concentrations was found. This confirms the participation of non-enzymatic oxidative decarboxylation of alpha-keto acids in the hydrogen peroxide decomposition process.