Horseradish peroxidase-mediated in situ synthesis of silver nanoparticles: application for sensing of mercury

Abstract

An indirect enzyme-induced in situ formation of silver nanoparticles (AgNPs) was investigated for the spectrophotometric detection of inorganic mercury (Hg²⁺) in aqueous solution. The enzymatic activity of horseradish peroxidase (HRP) assists the formation of AgNPs, which is prevented upon the addition of an excess amount of hydrogen peroxide (H₂O₂). On addition of the enzyme substrate 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt (ABTS), the excess amount of H₂O₂ converts the ABTS into the radical cation (ABTS˙⁺), which could help in restarting the HRP activity for the formation of AgNPs. The addition of increasing amounts of Hg²⁺ (HRP inhibitor) in the reaction mixture resulted in the progressive inhibition of the enzyme activity and could cause AgNP accumulation. The morphological changes caused during the enzyme-induced formation of AgNPs were further studied by transmission electron microscopy (TEM). The developed probe achieved a good limit of detection 0.78 × 10⁻⁹ M and a limit of quantification of 2.6 × 10⁻⁹ M in aqueous solution. The probe also demonstrated high recovery percentages for Hg²⁺ spiked in environmental matrices, highlighting the importance of the developed probe.