Electrochemical detection of extracellular hydrogen peroxide in Arabidopsis thaliana: a real‐time marker of oxidative stress

ABSTRACT An electrochemical approach to directly measure the dynamic process of H2O2 release from cultures of Arabidopsis thaliana cells is reported. This approach is based on H2O2 oxidation on a Pt electrode in conjunction with continuous measurement of sample pH. For [H2O2] 1 mm), the amperometric response can be described by Michaelian‐type kinetics and a mathematical expression relating current intensity and pH was obtained to quantitatively determine H2O2 concentration. At pH 5.5, the detection limit of the sensor was 3.1 µm (S/N = 3), with a response sensitivity of 0.16 Am−1 cm−2 and reproducibility was within 6.1% in the range 1–5 × 10−3 m (n = 5). Cell suspensions under normal physiological conditions had a pH between 5.5–5.7 and H2O2 concentrations in the range 7.0–20.5 µm (n = 5). The addition of exogenous H2O2, as well as other potential stress stimuli, was made to the cells and the change in H2O2 concentration was monitored. This real‐time quantitative H2O2 analysis is a potential marker for the evaluation of oxidative stress in plant cell cultures. Evaluation of H2O2 content in extracellular media can be an important oxidative burst marker in plant cell cultures. This paper shows an electrochemical method based on a Pt‐electrode which can be used for real‐time course analysis of H2O2 concentrations in plant cell suspensions. The protocol has been satisfactory applied in Arabidopsis thaliana cell suspensions and could be a first step in the development of future electrodes for in vivo monitoring of H2O2.