Cr(III)(picolinate)3 [Cr(III)(pic)3] is currently used as a nutritional supplement and for treating Type-2 diabetes. The effect of Cr(III)(pic)3 uptake in peripheral blood lymphocytes is investigated in this study. From the cytotoxicity data, DNA fragmentation pattern, Annexin V staining, TUNEL positivity and the ultrastructural characteristics such as chromatin condensation and formation of apoptotic bodies, it is clear that Cr(III)(pic)3 induces a concentration dependent apoptosis. It is shown that reactive oxygen species (ROS) produced by treatment with Cr(III)(pic)3 leads to apoptosis, since we find that pretreatment with N-acetyl cysteine inhibits the process. Using Western blotting technique and fluorescence measurements, the downstream signaling molecules have also been identified. Cr(III)(pic)3 treatment leads to collapse of the mitochondrial membrane potential, Bax expression, increase in cytosolic cytochrome c content and active caspase-3 and DNA fragmentation and all these manifestations are reduced by pretreating the lymphocytes with N-acetyl cysteine. Thus, it is shown that Cr(III)(pic)3 is cytotoxic to lymphocytes with ROS and mitochondrial events playing a role in bringing about apoptosis.