Detection of Cu²⁺ Using Methanobactin-coupled Lipase Biosensor with Differential Pulse Voltammetry

In this study, a methanobactin (Mb)-coupled lipase biosensor was constructed based on the fact that Mb can specifically capture Cu²⁺. Lipase immobilized on gold nanoparticles (Lipase@AuNPs) was successfully prepared and its structure and morphology were characterized by fluorescence, ultraviolet-visible (UV-vis) and Fourier transform infrared spectroscopy (FTIR) and scanning transmission electron microscopy (STEM). The biosensor could monitor the response of electrical signals during the hydrolysis of triolein by lipase. Cu²⁺ specifically combined with Mb and was enriched around lipase, which inhibited the catalytic activity of lipase and significantly reduced the current intensity, thus allowing rapid qualitative and ultra-trace reverse quantitative detection of Cu²⁺. The results showed that the optimal detection conditions, as determined by differential pulse voltammetry (DPV), were as follows: substrate concentration 2 g/100 mL and buffer pH 7.5. The sensor current difference had a good linear relationship with Cu²⁺ concentration in the range of 1–100 nmol/L. The regression equation was as follows: y = 0.228x + 0.7748, with a correlation coefficient (R²) of 0.9950, and the detection limit of this method was 0.03 nmol/L (R_SN = 3). Therefore, the new lipase biosensor has high sensitivity, specificity and stability, which lays a research foundation for the detection of trace and ultra-trace heavy metals in foods.