We investigated peroxide and superoxide accumulation, cytochrome c nature and release from mitochondria, as well as caspase activation during exposure of HL-60 cells to H 2 O 2 and the protective effect of ascorbic acid. Exposure of the cells to 100μM H 2 O 2 resulted in intracellular accumulation of peroxides, denaturation of cytochrome c that was identified in the mitochondria and cytosol, release of native cytochrome c to the cytosol and fall in mitochondrial membrane potential (ΔΨ m ). Loading of cells with ascorbic acid before exposure to H 2 O 2 resulted in a dose-dependent protective effect against: intracellular accumulation of peroxides, ΔΨ m alteration, cytochrome c denaturation and release. The accumulation of peroxides induced processings and activations of procaspase-8, -9 and -3. Double staining with antiserum which recognizes the p18 subunit of cleaved caspase-3 and with Hoechst had shown that a high percentage of cells exposed to 100μM H 2 O 2 stained positively with the antibody and showed features of apoptosis. Ascorbic acid loading prevented caspase activation induced by H 2 O 2 . We conclude that ascorbic acid protects against activation of apoptotic cascades in HL-60 cells exposed to H 2 O 2 and against apoptosis.