Development of glycerol biosensor based on co-immobilization of enzyme nanoparticles onto graphene oxide nanoparticles decorated pencil graphite electrode.

An improved amperometric biosensor was fabricated by immobilizing glycerol kinase (GK) and glycerol-3-phosphate oxidase (GPO) nanoparticles (NPs) onto graphene oxide nanoparticles (GrONPs) modified pencil graphite (PG) electrode. The GKNPs, GPONPs and GrONPs were characterized by UV spectroscopy, and transmission electron microscopy (TEM). The working electrode (GKNPs/GPONPs/GrONPs/PGE) was characterized by scanning electron microscopy (SEM), electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) techniques. The biosensor exhibited optimal current response at an applied potential of 0.45 V, pH 8.0, and 35 °C. The biosensor displayed a wide linear response for glycerol concentration from 0.001 to 60 mM with a detection limit of 0.002 μM. Moreover, a very high sensitivity 121.45 μA·mM-1·cm-2, rapid response time (2 s) and a good concurrence with the standard enzymic colorimetric technique with a correlation coefficient (R2 = 0.99) was offered by the present biosensor. Evidently, biosensor revealed an analytical recovery of 98.5% after addition of glycerol to the sera samples. Within and between batches studies of working electrode demonstrated coefficients of variation of 0.098% and 0.101%, respectively. The biosensor measured blood serum glycerol level in patients suffering from hyperglyceridemia. The biosensor lost 25% of its initial activity after its regular use over a period of 210 days, at 4 °C storage condition.