DNA Strand Breaks by Metal‐Induced Oxygen Radicals in Purified Salmonella typhimurium DNA

 Purified Salmonella typhimurium DNA was incubated for 1h at 37°C with various concentrations (10–100 μM) of transition metal ions (Fe 2+ , Fe 3+ , Cu 2+ , Ni 2+ , Cd 2+ ), with various concentrations (0.1–100 mM) of H 2 O 2 , and with various concentrations of each transition metal ion in the presence of various concentrations of H 2 O 2 . Damage to DNA was assessed by electrophoresis of the reaction mixtures in 1% agarose gel. Breakage of the DNA strands would produce a series of DNA fragments resulting in a smear in the gel, while intact DNA produced a single band. Results showed that no damage to the DNA was detectable after incubation with either H 2 O 2 alone or either of the metal ions alone. However, all of the metal ions investigated triggered DNA breakage in the presence of H 2 O 2 . The extent of breakage depended on the metal ion and on its concentration, as well as on the H 2 O 2 concentration. Addition of either EDTA or catalase to the reaction mixture completely inhibited the DNA degradation, confirming the involvement of both the metal ion and the H 2 O 2 in the breakage of DNA strands. Production of the hydroxyl radical when H 2 O 2 and a metal ion were both present in the reaction mixture was evidenced by the thiobarbituric acid method. The most extensive damage was caused by Cu 2+ followed, in decreasing order, by Fe 2+ , Fe 3+ , Ni 2+ , and Cd 2+ .