Toluidine blue adsorbed on alcohol dehydrogenase modified glassy carbon electrode for voltammetric determination of ethanol

A novel toluidine blue O (TBO) adsorbed alcohol dehydrogenase (ADH) biocomposite film have been prepared through simple adsorption technique with the help of electrostatic interaction between oppositely charged layers. Nafion (NF) coating was made on top of the biocomposite film modified glassy carbon electrode (GCE) to protect ADH from leaching. The fabricated ADH/TBO/NF biocomposite electrode remains highly stable in the pH range from 4 to 13. More facile electron transfer process occurs at ADH/TBO/NF biocomposite than at TBO/NF film, which is obvious from the six folds increase in ks value. Maximum surface coverage concentration (Γ) of TBO is noticed at ADH/TBO/NF film, which is 82% higher than at TBO/NF and 15% higher than at ADH/TBO film modified GCEs. Electrochemical impedance spectroscopy studies reveal that ADH has been well immobilized in the biocomposite film. Scanning electron microscopy studies confirm the discriminate surface morphology of various components present in the biocomposite film. Cyclic voltammetry studies validate that ADH/TBO/NF biocomposite film exhibits excellent electrocatalytic activity for ethanol oxidation at low over potential (Ipa = −0.14 V). The same studies show biocomposite film possesses a good sensitivity of 7.91 μA M−1 cm−2 for ethanol determination. This above sensitivity value is 17.40% higher than the sensitivity obtained for TBO/NF film (6.74 μA M−1 cm−2). Further, using differential pulse voltammetry, a sensitivity of 1.70 μA M−1 cm−2 has been achieved for ADH/TBO/NF biocomposite film.