A biosensor based on cytochrome c immobilization on apoly-3-methylthiophene/multi-walled carbon nanotubes hybrid-modified electrode. Application to the electrochemical determination of nitrite

摘要

A cytochrome c (Cyt c) electrochemical biosensor constructed by immobilization of the biomolecule on a hybrid material consisting of the conducting polymer poly-(3-methylthiophene) (P3MT) and multiwalled carbon nanotubes (MWCNT) is reported. Cyt c was immobilized onto 1-cysteine (i-Cys) self-assembled monolayer-modified P3MT/MWCNT/GCE at neutral pH, where positively charged Cyt c strongly interacts with 1-Cys negatively charged sites. The electron transfer rate constant, ks, was 0.49 ± 0.01 s~(.a(xm-c--, n1Udn. dt6heo1er1, ts1h±luer 0 2 foa0cpcte.i1~F mmc8oizv)edr(a)ge). of Cyt conditions, the mediatorless biosensor exhibits good electrocatalytic activity towards H 2O2 reduction. The amperometric measurements of H202 at an applied potential of 0.0 V in 0.05 M Tris-HCI buffer of pH 8.0 exhibited good repeatability (RSD = 3.1%, n = 10) and an acceptable reproducibility for the biosensor preparation (RSD = 8.1%, n = 5). The lifetime of the biosensor was evaluated. A linear calibration graph for H 2O2 (r = 0.998) was obtained between 0.7 and 400 p.M, with a detection limit of 0.23 p.M. The analytical characteristics for H 202 determination were found to be better than those reported for other nanostructured cytochrome c biosensors. The apparent Michaelis-Menten constant for the reaction between Cyt c and H202 was 451 pM. The developed Cyt c/I-Cys/P3MT/MWCNT/GCE biosensor was applied for the biocatalytic determination of nitrite through the oxidation of NOi to NOi by electrogenerated [Fe4*-Cyt c] species. Amperometry in stirred PBS solutions at a potential of +0.9 V allowed nitrite to be determined in a linear range between 10 and 100 pM, with a detection limit of 0.5 pM.