Enzyme-immobilized graphene oxide-based electrochemical biosensor for glutathione detection

Glutathione acts as a natural antioxidant in the human body and the reduction of its content is a sign of oxidative stress. In this study, a sensitive electrochemical sensor was developed using laccase enzyme immobilized onto graphene oxide (GO) for detection of glutathione. The surface of the indium tin oxide (ITO) was modified with GO via a drop casting method. Subsequently, laccase was immobilized onto the modified ITO decorated with GO. The modified electrode was characterized using field-emission scanning electron microscopy (FESEM), X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The FTIR spectra of laccase/GO confirmed the successful immobilization of laccase onto GO sheets. FESEM analysis revealed the transformation from a layered, wrinkled structure to a compact, smooth surface with spherical laccase, confirming successful enzyme integration. Raman analysis confirmed successful laccase immobilization onto GO, as evidenced by structural changes in the D and G bands, highlighting the modification of the material. The cyclic voltammetry measurements revealed that laccase/GO/ITO exhibited better electrocatalytic activity toward oxidation of GSH in acetate buffer solution than the bare ITO electrode. This newly developed electrode exhibited a good response to glutathione with a wide linear range from 1 μM to 100 μM, a limit of detection of 0.89 μM and high sensitivity (6.51 μA μM⁻¹). Furthermore, it exhibited excellent selectivity, repeatability, and long-term stability. The modified electrode was successfully used for the detection of GSH in a real sample, offering satisfactory results.