*3.11. Determination of Reactive Oxygen Species (ROS)*

The H2O2 levels were determined by the cell permeant probe 2 -7 -dichlorodihydrofluorescin diacetate (H2DCFDA). Briefly, after 24, 48 and 72 h of exposure to specific red onion sample concentrations (0, 0.5, 1 and 5 mg/mL), based on the results of the cell viability assay, the media were changed and oxidative stress was induced by 50 μM tertbutyl hydroperoxide (T-BHP) treatment (Sigma-Aldrich, B2633, St. Louis, MO, USA), as described in [53,54]. After 45 min of treatment, the cells were incubated in the dark, at 37 ◦C for 20 min with 10 μM H2DCFDA. After that, the cells were washed and resuspended in 150 μL of PBS, and the H2O2-dependent oxidation of the fluorescent probe was measured by the Victor 2030 multilabel reader (PerkinElmer, Waltham, MA, USA) (at 507-nm excitation and 530-nm emission wavelength).
