Fabrication of the carbon paste electrode modified with Trametes versicolor laccase immobilized on carboxyl functionalized multi-walled carbon nanotubes and its application for measurement of dopamine.

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Abstract

Dopamine (DA) shows numerous roles in a wide range of physiological and pathological processes. In this study, an immobilized laccase-derived biosensor was developed for DA detection. The carboxyl functionalized multi-walled carbon nanotubes (MWCNTs-COOH) was applied for immobilization of laccase from Trametes versicolor (TvLac). According to Plackett-Burman statistical design, the optimum conditions showed at 5 mg/mL of MWCNTs-COOH, 25 mM phosphate buffer (pH 6.0), sonication time for 15 min, 2.5 U/mg of enzyme concentration, immobilization time for 4 h at 4 °C, and rotation at 100 rpm. At these conditions, the experimental and predicted specific activities were 14.19 ± 1.41 U/mg and 13.99 ± 1.54 U/mg, respectively. The activity of immobilized TvLac was >90 % at 60 °C and pH 7.0 as well as after 10 sets of uses. The carbon paste electrode (CPE) modified with the immobilized TvLac was then fabricated, characterized and applied as a biosensor (TvLac@MWCNTs-COOH/CPE) for determination of DA. The mean of diffusion coefficient for DA was considered to be 9.1 × 10-6 cm2/s. The TvLac@MWCNTs-COOH/CPE represented a linear dynamic range of 0.005-100.0 μM with detection limit of 1.0 nM. The TvLac@MWCNTs-COOH/CPE might be introduced as a suitable sensor for monitoring of DA in real specimens which merit further studies.