Occupational exposure in a copper smelting industry may produce various adverse health effects including cancer. Despite a number of well-documented studies reporting an increased risk of cancer among copper smelter workers, the data on genotoxic effects in this industry are scarce. In view of the above, an assessment of DNA damage in peripheral blood leukocytes by Comet assay from copper smelter workers was undertaken. Additionally, the proton-induced X-ray emission (PIXE) analysis was assessed to determine the metal content of samples. The study was conducted with all workers from a copper smelter (males; n = 11), and a control group (n = 11) was recruited. The results of our study showed a significant increase (p < 0.001; Mann-Whitney test) in DNA damage in peripheral blood lymphocytes of smelter workers, compared to the controls (p < 0.001; Mann-Whitney test). No correlation between DNA damage or metal concentration and age mean or time of exposure was found under study. Our findings indicate that copper smelter workers have increased levels of DNA damage in somatic cells, suggesting a potential health risk for the workers. PIXE results show the presence of copper, iron, and other metals.