The influence of substrates addition order on colorimetric assay based on MnO₂ nanocubes: A novel turn-off H₂O₂ assay strategy in water-soak foods

Herein, novel MnO₂ nanocubes were facilely synthesized by manipulating the dosage of KMnO₄ and L-Dopa with the aid of ultrasound. The as-prepared MnO₂ nanocubes exhibited interesting oxidation activity which was influenced by the addition order of substrates (H₂O₂ and 3,3,5,5-tetramethylbenzidine (TMB)). After the mechanism study, we found that H₂O₂ and TMB can be competitively oxidized by MnO₂ nanocubes, which was different from the peroxidase- and oxidase-like activities. According to the discovery, a novel turn-off H₂O₂ assay method based on MnO₂ nanocubes was established, where H₂O₂ was firstly incubated with MnO₂ nanocubes for 3 min and then TMB was added for the instantaneous chromogenic reaction. In addition to the shorter operation time, the colorimetric results were less affected by temperature and unchanged within 30 min without terminating reaction. Moreover, the method showed ultra-high sensitivity with low limit of detection (0.027 μmol L⁻¹) and acceptable reliability for H₂O₂ assay in water-soak foods.