Abstract Nitroxyl (HNO/NO−) is a molecule which results from one-electron reduction of nitric oxide (NO). It is considered to be a pharmacologically important particle because of its cardiovascular effects and potential anticancer activity. Due to molecule instability studies on nitroxyl biological activity require the use of donor compounds. In the present study Angeli’s salt (Na2N2O3) was used as the nitroxyl donor and cytotoxicity of HNO/NO− against human lymphocytes and A549 and HT29 cancer cells was examined. The studies were performed under different pH conditions (pH 6.2 and 7.4) because it was suggested that formation of HNO and its toxicological effects may be stronger in tissues subjected to acidosis such as cancerous ones. It was shown that nitroxyl exerted cytotoxic effect against all three types of cells. Nitroxyl induced both apoptotic and necrotic cell death. The cytotoxic effect analyzed directly after cell treatment was stronger than that observed 24h later. Differences in cell sensitivity to nitroxyl were observed – proliferating lymphocytes were the most sensitive cells. It was also shown that pro-oxidant activity of nitroxyl was stronger under slightly acidic conditions as compared to physiological pH but this difference was not always reflected in the observed cytotoxic effect, especially when the effect was measured 24h after cell treatment. Thus, relatively high toxicity of nitroxyl against normal cells and its ability to induce not only apoptotic but also necrotic cell death make use of this molecule in cancer therapy questionable.