2.3. Enzymatic activity assay

Laccase activity was assayed at room temperature by monitoring the oxidation of ABTS at 420 nm (ε₄₂₀ 36×10³ M⁻¹ cm⁻¹). The assay mixture contained 2×10⁻³ M ABTS in 0.1 M sodium citrate buffer, pH 3.0. Laccase activity towards 2,6‐dimethoxyphenol (DMP) was assayed in a mixture containing 1×10⁻³ M DMP in McIlvaine's citrate phosphate buffer adjusted to pH 5.0. Oxidation of DMP was followed by an absorbance increase at 477 nm (ε₄₇₇ 14×10³ M⁻¹ cm⁻¹). One unit of activity is defined as the amount of laccase oxidizing 1 µmol of substrate per min. Laccase activity towards aniline was monitored determining the Abs_410nm after 30 min of incubation of 300 mM aniline with 1 U/mL of enzyme at 28°C. Different enzymes were tested: the high‐redox potential T. versicolor laccase B, TvB ²³, ²⁷, P. ostreatus, POXC (E° 0.74, pH 5) ²⁸, rPOXA1b (E° 0.65, pH 6) ²⁶ and its improved variant 1H6C (E° 0.77, pH 6) ²⁹.