Laccase Immobilization on Carbon-Based Materials Derived from Spent Brewery Grains: Optimization and Stability Evaluation.

From the paper

Abstract

Enzyme immobilization onto solid supports enhances their stability, reusability, and efficiency. This work investigates the physical immobilization of laccase (Lac) from Trametes versicolor (purchased, EC 1.10.3.2, ≥0.5 U/mg) onto two carbon-based materials: activated carbon (AC) and biochar (BC), obtained from spent brewery grains (SBGs) through microwave pyrolysis (with and without chemical activation, respectively), generating SBG-AC/Lac and SBG-BC/Lac. Various immobilization conditions (pH 3.5-6.5, Lac concentration 1-10 mg/mL) were tested, with immobilization up to 80 ± 6% (for Lac 1 mg/mL, pH 5.0 in SBG-AC/Lac) and maximum activities of 5.5 ± 0.2 U/g (SBG-AC/Lac) and 4.6 ± 0.5 U/g (SBG-BC/Lac) at pH 3.5 and 40 °C. Although SBG-AC led to a higher immobilization %, SBG-BC was a greener alternative, requiring no chemical activation during production. Kinetics analysis with a typical Lac chromogenic substrate revealed higher values of KM (Michaelis constant) for SBG-BC/Lac compared with free Lac (Lacf) (indicating lower substrate affinity), but higher stability, retaining ~60% activity after 24 h, while Lacf was nearly inactive. These results demonstrate the potential of SBG-BC as a sustainable support for Lac immobilization in applications such as wastewater treatment and environmental monitoring.