RESUMO
An improved cytochrome c (Cyt c) biosensor based on immobilization of cytochrome c oxidase (COx) on the surface of graphene oxide nanoparticles (GONPs) electrodeposited onto pencil graphite (PG) electrode. Characterization of graphene oxide nanoparticle was done by Transmission electron microscopy (TEM), Fourier transform infra-red spectroscopy (FTIR) and X-ray diffraction study (XRD). The working electrode (COx/GONPs/PG) was characterized at its different stages of fabrication by scanning electron microscopy (SEM) and FTIR. Fabrication of Cyt c biosensor was done by connecting COx/GONPs/PG as working electrode, Ag/AgCl as reference electrode and Pt as auxiliary electrode to potentiostat. The mechanism of detection of present biosensor was based on oxidation of Cyt c (reduced) to Cyt c (oxidized) by COx resulting in flow of electrons through GONPs to the PG electrode, hence current generated is proportional to the concentration of Cyt c. Present biosensor exhibited optimum potential at 0.49 V with optimum pH 7.5 and optimum temperature 35°C. Biosensor showed linearity within 40-180 ng/ml having 40 ng/ml limit of detection. The precision i.e. within and between-batch coefficients of variation (CVs) were found <0.04% and <0.21% respectively. The enzyme electrode lost 50% of its initial activity when operated for more than 6 months on weekly basis. It was applied for detection of Cyt c level in in apparently healthy and diseased human sera. The present biosensing method was co-related with standard colorimetric method and co-relation coefficient was found 0.99.
