Methods to assess free radicals and oxidative stress in biological systems

Oxidative stress results from a disruption of the prooxidant/antioxidant ce llular balance and monitoring free radical status becomes an interesting ch allenge in animal and human nutrition. In the present work, merits and limi tations of different analytical techniques (HPLC, GC-MS, fluorometric and c olourometric assays, ELISA, gel electrophoresis) for the measurement of rad ical mediated alterations in the cellular integrity of lipids (malondialdeh yde, hydrocarbon gases, F2-isoprostanes) proteins (protein carbonyls, 3-nit rotyrosine) and DNA (8-hydroxy-2'-deoxyguanosine) are discussed. Besides th ese indirect methods, owing to the fact that free radicals are paramagnetic , electron paramagnetic resonance spectroscopy combined with spin trapping has become a valuable tool to directly assess and to better understand the mechanisms of free radical reactions. With this approach a radical that is too short-lived to be detected, adds to a spin-trapping agent to form a rel atively long-lived radical adduct. Information obtained from the hyperfine splitting of the spin-trapped adduct can provide identification and quantif ication of the originally generated free radicals.