Peroxidase-mimetic colloidal nanozyme from ozone-oxidized lignocellulosic biomass for biosensing of H2O2 and bacterial contamination in water

Nanozymes, possessing enzyme-like traits, have gained tremendous attention for their functionality, ease of production, economical synthesis, and stability. Majority of reported nanozymes in literature, for analyte detection are metal-based compounds, transition metal dichalcogenides or single-atom nanozymes. In this study, we report for the first time, a novel peroxidase-mimic, colloidal dendritic nanozyme from lignin-rich agro-industrial residue (coconut husk) by ozonolysis. Synthesized nanozyme exhibited peroxidase-mimic activity in sensing H₂O₂, with a wide range of substrates and detection techniques. When 3,3′,5,5′-tetramethylbenzidine (TMB) and 2′,7′–dichlorofluorescin diacetate (DCFDA) were used, the nanozyme demonstrated ultrafast kinetic behaviour with LOD of 43.60 ± 2.41 µM and 1.25 ± 0.31 µM H₂O₂, by colorimetric and fluorimetric assays, respectively. The nanozyme-based H₂O₂ sensing platform, was further utilized for detection of pathogenic bacteria namely Escherichia coli, Listeria monocytogenes, Staphylococcus aureus and Pseudomonas aeruginosa, and for total bacterial load in water. Notably, it demonstrated high sensitivity in the detection of P. aeruginosa with LOD as low as 7 CFU/mL with both fluorimetric and electrochemical methods. Ultrasensitive detection of total bacterial load could also be achieved with 5.5 × 10² CFU/mL, 5.5 × 10¹ CFU/mL, and 4.1 × 10¹ CFU/mL by colorimetric, fluorometric, and electrochemical techniques, respectively. Results of the study thus indicate, that the developed nanozyme-based sensing platform had high sensitivity for detection of bacteria as well as versatility with diverse analytical approaches enabling potential practical application for “onsite” monitoring of water quality, especially in rural settings. This biological mimic can also be used in sensor platforms where H₂O₂ is measured and applied for output signaling.