Development of amperometric biosensors using thiolated tetrathiafulvalene-derivatised self-assembled monolayer modified electrodes

The usefulness of tetrathiafulvalene (TTF) derivative self-assembled monolayers (SAMs) modified electrodes as platforms for the construction of electrochemical biosensors is evaluated by using a 4-mercaptotetrathiafulvalene (HSTTF) modified gold electrode for the preparation of a fructose electrochemical biosensor. The electroactive SAM enhances the electron transfer between the immobilized FDH and the electrode surface, the electrochemical oxidation of HSTTF being used to monitor fructose concentration. The amount of HSTTF employed to form the electroactive SAM (2.2 μg), the FDH loading (20.4 U), and the applied potential to the biosensor (+0.40 V) were optimized. The stability of the FDH-HSTTF-AuE was evaluated under several aspects, a low reproducibility between different biosensors being observed. Calculation of the apparent Michaelis–Menten constant indicated that the affinity of the enzyme for the substrate is practically non-affected by the immobilization procedure. A detection limit of 2.7 × 10−7 mol L−1 fructose was achieved. The reproducibility of the responses obtained with different biosensors was largely improved by forming mixed monolayers consisting of HSTTF and mercaptohexanol (MCH). The limit of detection calculated for the FDH-MCH/HSTTF-AuE biosensor is 4.2 × 10−7 mol L−1 fructose. The performance of this biosensor for the determination of fructose in real samples is evaluated by the analysis of a Cola soft drink sample.