Membrane separation assisted colorimetric/fluorescent detection of β-galactosidase-positive bacteria in milk and milk powder based on the oxidase-like activity of CoOOH nanosheets.

Species-specific enzymes provide a substantial boost to the precision and selectivity of identifying dairy products contaminated with foodborne pathogens, due to their specificity for target organisms. In this study, we developed cobalt oxyhydroxide nanosheets (CoOOH NSs) for a dual-mode biosensor capable of detecting β-galactosidase (β-Gal)-positive bacteria in milk and milk powder. The sensor exploits the oxidase-mimicking activity of CoOOH NSs, where β-Gal converts the substrate β-D-galactopyranoside to p-aminophenol, reducing CoOOH NSs to Co and inhibiting the formation of the blue product from 3,3',5,5'-tetramethylben-zidine. Sensitivity was enhanced through membrane filtration and β-Gal induction by isopropyl β-D-thiogalactoside. The assay achieved a detection limit of 5 cfu mL and demonstrated recoveries (90.7 % to 103 %) and relative standard deviations <5.7 % in milk and milk powder samples. These findings underscore the potential of the sensor for detecting β-Gal-positive bacteria in dairy products.