Production of a high level of laccase by submerged fermentation at 120-L scale of Cerrena unicolor C-139 grown on wheat bran.

Submerged fermentation in a stirred bioreactor of the white rot fungus Cerrena unicolor C-139 was done at a 120-L scale in the presence of wheat bran as a cheap lignocellulosic substrate for fungus growth and laccase production. Enzyme monitoring showed that laccase production started after 2 days of cultivation, attaining a maximum activity of 416.4 U·mL(-1) at day 12 of fermentation. After treatment of culture liquid by successive micro- and ultrafiltration (5kDa), a liquid concentrate containing 22203176 units of laccase was obtained. Obtaining large amount of laccase is essential for various industrial applications, including detoxification of industrial effluents, textile and petrochemical industries, polymer synthesis, bioremediation of contaminated area, stabilization of beverages, production of cosmetics, manufacture of anti-cancer drugs, and nanobiotechnology. The cultivation method and the fungal strain used here provided a substantial amount of enzyme produced at a price lower than 0.01 € cent/unit enzyme.

Immobilized laccase of Cerrena unicolor for elimination of endocrine disruptor micropollutants.

The white-rot fungus Cerrena unicolor C-139 produced 450 000 U l(-1) of laccase when cultivated in submerged (50 ml) fermentation of wheat bran. Laccase (benzenediol: oxygen oxidoreductase, EC 1.10.3.2.), from C. unicolor C-139 was immobilized covalently on control porosity carrier silica beads. The activity of the immobilized laccase was approximately 15.8 units per gram of silica beads. The pH optimum was between 2.5 and 3.0 for free and immobilized laccase. The immobilization of enzyme appeared to be the main factor for retention of laccase activity at high temperature of 80 °C. The apparent K(m) value (100 µmol) of immobilized laccase from C. unicolor C-139 was 6.7 times higher than free laccase (15 µmol) using 2,2-azino-bis-[3-ethylthiazoline-6-sulfonate] (ABTS) as the substrate. Immobilized laccase was able to eliminate 80 % of Bisphenol A, 40 % of Nonylphenol, and 60 % of Triclosan from solutions containing 50 µmol of each micropollutant separately. The experiments were run three times consecutively with the same immobilized laccase without loss of enzyme activity.