Determining Hydrogen Peroxide Content in Plant Tissue Extracts

Hydrogen peroxide (H2O2), like other reactive oxygen species (ROS), plays a dual role in physiology of plant organisms. On the one hand, ROS are capable of initiating oxidative stress, thus resulting in damage and death of the cells and whole plants. On the other hand, ROS are signaling molecules that promote the formation of adaptive mechanisms and improve plant resistance. Therefore, H2O2 is now primarily considered as a signaling molecule that acts as a key component of hormonal signaling networks as well as a regulator of growth, development, stomatal guard cell movements, and adaptive responses to biotic and abiotic stressors. The majority of the methods used to measure H2O2 and other ROS are being criticized for the lack of specificity and, most important, insufficient reliability. Herein, possible causes for variability of results were analyzed, general requirements for sample pretreatment were formulated, and criteria to check if reliable data are obtained were proposed. The authors outlined in detail the benefits and shortcomings of commonly used methods for assaying H2O2 concentration in plant extracts, including those based on the oxidation of Fe2+, I–, luminol, and substrates of peroxidase reaction as well as the methods relying on the complex formation between H2O2 and Ti(IV).