Spent grain as a sustainable and low-cost carrier for laccase immobilization

Spent grain is promising lignocellulosic by-product support for laccase immobilization. The waste digestion with two different approaches (HCl/NaOH and H₂SO₄/NaOH) was performed. Different procedures (soaking and dropping), based on chemical and physical reactions, were also used to obtain the highest immobilized activity. Results showed that H₂SO₄/NaOH digestion guaranteed an immobilized activity five times higher than HCl/NaOH digestion. The best immobilization conditions with physical dropping procedure resulted in the highest immobilized activity on digested spent grain (2500 U/Kg). Good reusability (42% of activity retained after four cycles), and lower catalytic efficiency (V_max/K_m) of 0.053 min^-1 than free laccase (0.14 min^-1) with ABTS as substrate, were also obtained. Besides, when 20 mg of biocatalyst (0.02 U) were tested for syringic acid removal, complete oxidation of the phenol was achieved in just 4 h.