THE JOURNAL OF BIOLOGICAL CHEMISTRY Detection of Lipid Radicals by Electron Paramagnetic Resonance Spin Trapping Using Intact Cells

Electron paramagnetic resonance (EPR) spin trap-ping was used to detect lipid-derived free radicals gen-erated by iron-induced oxidative stress in intact cells. Using the spin trap w(4-pyridyl 1-oxide)-N-tert-bu-tylnitrone (POBN), carbon-centered radical adducts were detected. These lipid-derived free radicals were formed during incubation of ferrous iron with U937 cells that were enriched with docosahexaenoic acid (22:6n-3). The EPR spectra exhibited apparent hyper-fine splittings characteristic of a POBN/alkyl radical, aN = 15.63 f 0.06 G and an = 2.66 & 0.03 G, generated as a result of &scission of alkoxyl radicals. Spin adduct formation depended on the FeSO, content of the incu-bation medium and the number of 226-enriched cells present; when the cells were enriched with oleic aci