Detection and Measurement of ROS in Tobacco Leaves

Leaf metabolism produces hydrogen peroxide (H₂O₂) at high rates, high level H₂O₂ accumulation can cause oxidative stress. This protocol describes a method for determining H₂O₂ concentration in tobacco leaves. In this method all extractions were performed with HClO₄, neutralized, and pretreated with ascorbate oxidase to eliminate ascorbate interferences. H₂O₂ content was determined using a colorimetric assay spiked with an internal control. Interfering peroxides were determined in parallel using a negative control treated with catalase and subsequently subtracted.