AP-22: One-Step Chemiluminescent Assay for Hydrogen Peroxide Analysis in Water

Abstract

In this work, we aim to develop a simple, and rapid, nonenzymatic and homogeneous biosensor for sensitive and rapid quantification of hydrogen peroxide.

Methods: In this technique, hemoglobin was used as a bioreceptor, where heme groups act as electroactive centers to catalyze hydrogen peroxide reduction. The chemiluminescence reagent; luminol is also a peroxidase substrate and can be oxidized by hemoglobin thus generating a CL signal. The working principle was based on the competition between hydrogen peroxide and luminol towards hemoglobin. A 96-well microplate was used to perform this assay. For thatat, 25 µL of Hb solution (3 µg mL−1) was added to each well, followed by 25 µL of different concentrations of H2O2. After incubation, 25 µL of luminol solution was also added. Then, the chemiluminescence intensity was measured at an emission wavelength of 425 nm.

Results and discussion: The detection principle is mainly based on the chemiluminescence signal diminution in the presence of H2O2. Because this latter will react with Hb that is essential for luminol reaction, thus leading to a lower chemiluminescence signal. AlsoUnder optimized conditions, the chemiluminescent signal decreased with increasing hydrogen peroxide concentrations within the linear range of 0.5 to 12 mM, with a correlation coefficient R2 of 0.99762. The limit of detection was calculated to be as low as 0.308 mM. The selectivity of the biosensor was successfully demonstrated against different interferents.

Conclusion: In this work, an homogenous chemiluminescent assay was developed using Hb as a bioreceptor. The developed strategy provides a one step, simple, and low-cost bioanalytical method which can be applied for the monitoring of hydrogen peroxide