A bienzymatic lactate biosensor with co-immobilization of horseradish peroxidase and lactate oxidase on sugarcane-derived three-dimensional carbon nanoballs cross-linked carbon aerogels

In this work, a novel bienzymatic lactate biosensor was developed by co-immobilization of lactate oxidase (LOD) and horseradish peroxidase (HRP) onto the screen-printed electrode (SPE) modified by the three-dimensional (3D) carbon nanoballs cross-linked carbon aerogels (3D-CNCAs), which is derived from the biomass of sugarcane. The 3D-CNCAs, with large surface area and porous structure, play a crucial role in facilitating direct electron transfer between the electroactive center of HRP and the biosensor electrode. The experimental parameters including the amount of 3D-CNCAs suspension, the ratio of the two enzymes, the buffer pH and the operation potential were optimized. The bienzymatic lactate biosensor exhibits linear current response for lactate in the concentration ranges of 1-1690 μM, with a detection limit of 0.23 μM (S/N = 3), which is superior to the one based on monotypic enzyme of LOD. The bienzymatic lactate biosensor also exhibits a high level of selectivity, reproducibility, repeatability and long-term stability (87% of the original value after storage for 30 days), and was further applied in lactate detection in diluted real samples of lactic acid cream and commercial beverage. The outstanding analytical performance of the bienzymatic lactate biosensor demonstrates the preponderance of the bienzymatic over the one based on monotypic enzyme, and holds broad application prospects in the future.