Generation of hydroxyl radicals from metal-loaded humic acids

Humic acids (HAs) are naturally occurring biopolymers that are ubiquitous in our environment. They are most commonly found in the soil, drinking water, and a variety of plants. Pharmacological and therapeutic studies involving humic acids have been reported to some extent. However, when certain transition metals are bound to humic acids, e.g., iron and copper, they can be harmful to biological organisms. For this study, humic acids were extracted from German, Irish, and New Hampshire soils that were selectively chosen because of their rich abundance in humic material. Each sample was treated at room temperature with 0.1 M ferric and cupric solutions for 48 h. The amount of iron and copper adsorbed by humic acid was accurately quantitated using atomic absorption spectroscopy. We further demonstrate that these metal-loaded humic acids can produce deleterious oxidizing species such as the hydroxyl radical (HO-) through the metal-driven Fenton reaction. Electron paramagnetic resonance (EPR) employing spin trapping techniques with 5,5;dimethylpyrroline N-oxide (DMPO) is used to confirm the generation of hydroxyl radicals. The DMPO-OH adduct with hyperfine splitting constants A(N) = A(H) = 14.9 G is observed upon the addition of exogenous hydrogen peroxide. The concentration of hydroxy[ radical was determined using 4-hydroxytempo (TEMPO-OH) as a spin standard. The presence of another oxidizing species, Fe=0(2+), is also proposed in the absence of hydrogen peroxide.