2.2. Measurement of lipid hydroperoxides

Plasma (10 vol%, unless otherwise specified) was oxidized with multiple oxidants at 37 °C in PBS (pH 7.4) under air in the absence and presence of antioxidant. Lipid hydroperoxides was measured from the fluorescence intensity by DPPP oxide, the excitation and emission wavelength being 351 and 380 nm respectively, with a microplate reader, Spectra Max M2 (Molecular Devices, Sunnyvale, CA) equipped with a thermostatted cell maintained at 37 °C under air as reported previously [41]. The oxidation was started by the addition of the respective oxidant into the PBS solution of plasma in the presence of DPPP and additives. DPPP and antioxidants were added as DMSO solution. The concentration of DMSO was kept to or less than 2.5 vol%.

Since the plasma obtained from different mice contained different composition of lipids and antioxidants, the same plasma was used for a set of experiments. The experiments were repeated at least twice, in most cases more than three times, and the reproducibility was satisfactory.