A single step electrochemical integration of gold nanoparticles, cholesterol oxidase, cholesterol esterase and mediator with polypyrrole films for fabrication of free and total cholesterol nanobiosensors

The development of free and total cholesterol nanobiosensors based on a single step electrochemical integration of gold nanoparticles (AuNPs), cholesterol oxidase (COx), cholesterol esterase (CE) and a mediator with polypyrrole (PPy) films is described. The incorporation of the various components in the PPy films was confirmed by chronopotentiometry, cyclic voltammetry (CV), scanning electron microscopy, energy dispersive X-ray analysis (SEM-EDX), and Fourier transformed infrared (FTIR) spectroscopy. The free cholesterol, PPy-NO3--Fe(CN)64--AuNPs-COx, nanobiosensor achieved a minimum detectable concentration of 5 μM, a linear concentration range of 5–25 μM and a sensitivity of 1.6 µA cm−2 µM−1 in 0.05 M phosphate buffer (pH 7.00). For the total cholesterol, PPy-NO3--Fe(CN)64--AuNPs-COx-CE, nanobiosensor which also involved the co-incorporation of cholesterol esterase (CE) with the other components, the achieved performances include a minimum detectable total cholesterol concentration of 25 μM, a broader linear concentration range of 25–170 μM and a lower sensitivity of 0.1 µA µM−1 cm−2. Owing to its high selectivity, the presence of common interferants did not affect the total cholesterol measurement with the PPy-NO3--Fe(CN)64--AuNPs-COx-CE nanobiosensor. Both nanobiosensors were successfully used for direct and indirect determination of total cholesterol in human blood serum samples. [Display omitted] •Single step integration of AuNPs, two enzymes and mediator in a PPy Film.•Amperometric detection of free and total cholesterol.•Interference-free detection in presence of potential interferants.•Direct and indirect determination of total cholesterol in human serum samples.