Construction of a laccase mimic enzyme with fluorescence properties for kanamycin multi-mode analysis

Abstract

A copper-based dual-ligand biomimetic enzyme (Cu-BH) inspired by natural laccase was constructed, which had promising laccase-like activity and excellent fluorescence properties. More remarkably, Cu-BH can oxidize not only traditional phenolic substrates, but also catecholamines and amine substrates. As expected, OPD could be catalyzed by Cu-BH to produce the yellow fluorescent product DAP with absorbance and emission peaks at 426 nm and 566 nm, respectively. The generated DAP could quench the Cu-BH fluorescence at 440 nm by fluorescence internal filtration effect (IFE), thus constructing a ratiometric fluorescence signal(F566/F440). Kanamycin further promoted the catalytic activity of Cu-BH by accelerating electron transfer, resulting in a gradual increase in absorbance at 426 nm and fluorescence signal amplification at 566 nm. Based on this, a sensitive dual-mode sensing strategy was established to analyze kanamycin with the LOD of 0.033 and 0.26 nM, which was successfully applied to the monitoring of kanamycin in tap water, milk and honey. The visualization analysis was also achieved by RGB reading from a smartphone. In conclusion, this novel biomimetic enzyme (Cu-BH) had provided a new application path in bioanalysis and environmental monitoring.