Electroporation is a most popular method of cell membrane permeabilization, by pulsed electric fields. It allows foreign molecules to enter the cell and has been used for many biotechnological applications, including transformation of mammalian cells and plant protoplasts by exogenous genetic material. However, the mechanism underlying membrane electropermeabilization is still largely unknown. Evidence is presented here that electroporation under conditions compatible with cell survival induces lipid hydroperoxide formation in the membranes of animal and plant cells. Exposure to electric fields also enhanced up to 5-fold the spontaneous emission of light from both cell types, which paralleled the amount of conjugated hydroperoxides detected in cell membranes. The emitted photons were mainly in the red edge of the spectrum, suggesting the involvement of singlet oxygen. The presence of antioxidants during electroporation did not reduce the formation of hydroperoxides nor the permeability but quenched the luminescence.