In-situ synthesis of 2D nanozymes-coated cellulose nanofibers on paper-based chips for portable detection of biothiols

Background: Simple, fast and low-cost paper-based analytical devices (PADs) have a good application prospect for point-of-care detection of GSH. However, effective immobilization of functional nanomaterials onto cellulose, as a critical factor in the construction of PADs, presents numerous difficulties and challenges. Results: In this study, we have developed an exceptionally straightforward and environmentally friendly synthetic approach by using ovalbumin (OVA) as a bio-mineralization template for the preparation of MnO2 nanosheets. The MnO2 nanosheets produced in the solution phase exhibited excellent intrinsic nano-enzyme activity and biodegradability. The OVA-MnO2 nanosheets can effectively oxidize Amplex red in the absence of H2O2, enabling sensitive detection of GSH with a linear range of 5 nM-10 mu M and a detection limit as low as 2.8 nM. Furthermore, we utilized this method to facilitate in situ synthesis of OVA-MnO2 nanosheets directly on paper substrates. This approach eliminates the need for conventional stirring and centrifugation steps, greatly simplifying the fabrication process while reducing material usage and time expenditure. Characterization of the chemical composition and morphology confirmed the intimate growth of the 2D nano-enzymes on the cellulose fibers. Utilizing smartphone capabilities, the OVA-MnO2 nanosheet-modified PAD enabled instrument-free detection of GSH, demonstrating high sensitivity (0.74 mu M) and a wide linear response range (1-1000 mu M). Significance: The synthesis of MnO2 nanosheets directly on cellulose substrates substantially streamlines the modification workflow of PADs and reduces detection costs, offering new avenues for clinical diagnostics of relevant diseases.