It has been found previously that hydroxycobalamine (vitamin B12b) amplifies significantly the cytotoxic effect of ascorbic acid (vitamin C) added to cells for small a, Cyrillic long period of time (48 h). However, according to pharmacokinetics, the concentration of vitamin C in vivo decreases to a physiological value within a short period of time (2-3 h) after the injection. Therefore, in this study we examined the cytotoxic effect of a short-time (up to 2 h) exposure of human larynx carcinoma HEp-2 cells to a combination of vitamins B12b and C (B12b+C). The kinetics of the B12b+C-caused extracellular oxidative burst in this time interval was also explored. Vitamin B12b combined with ascorbic acid provoked a rapid accumulation of extracellular hydrogen peroxide followed by intracellular oxidative stress, DNA single-strand breaks, and the initiation of apoptosis. The chelators of iron phenanthroline and desferrioxamine prevented B12b+C-induced DNA single-strand breaks and cell death but not the accumulation of H2O2 in culture medium. The nonthiol antioxidants pyruvate and catalase were effective in preventing the prooxidant and cytotoxic effects of B12b+C. Thiols, when added simultaneously with the combined vitamins, inhibited these effects only partially (N-acetylcysteine, GSH) or even amplified them (dithiothreitol). The results obtained point to the determining role of oxidative burst and iron-dependent DNA damage in the cytotoxic effect of short-time exposure to B12b+C combination.