Synthesis of 8‐nitroguanine in calf thymus DNA by a Fenton‐like reaction

Abstract

8-Nitroguanine has been found in DNA and RNA. Its production in nucleic acids has been linked to inflammation, chronic hepatitis C, gouty arthritis, and premalignant lesions. Presumably in vivo, 8-nitroguanine is formed by the reaction of peroxynitrite, itself formed by the reaction of nitric oxide with superoxide produced by macrophages, with guanine. Recently, nitro-tyrosine was produced in vitro by a Fenton type reaction. We have found that 8-nitroguanine can also be produced in vitro by a Fenton type reaction. Calf thymus DNA was dissolved in 5mM phosphate buffer, pH 7.01, and treated with hydrogen peroxide, sodium nitrite, and various iron (II) and iron (III) salts. The solutions were incubated with shaking at 37°C and aliquots of hydrogen peroxide and sodium nitrite added. The solutions were processed to isolate 8-nitroguanine, which depurinates, and analyzed by HPLC using a C18 column and a photo-diode array detector. A compound with a retention time and spectra matching that of an 8-nitroguanine standard was found. The iron salts were not equally adept at producing 8-nitroguanine; this could be due to solubility or some unknown effect caused by the salts’ anion. It is possible that production of some 8-nitroguanine in vivo maybe due to Fenton like reactions.