Enzyme production by Trichoderma reesei Rut C-30 followed by enzymatic hydrolysis of different lignocellulosic materials

The filamentous fungus Trichoderma reesei is one of the main sources for cellulose degrading enzymes. We study the enzyme profile produced during the fungal growth on cellulosic and lignocellulosic substrates and their capacity to hydrolyze cellulosic and lignocellulosic substrates with different chemical and physical properties. The results will bring insight into the bottlenecks of enzymatic hydrolysis. During the enzyme production study, we grew T. reesei strain Rut C-30 in submerged fermentations on Avicel PH101, commercial cellulose, and industrial-like lignocellulosic substrates from spruce. These substrates were produced during the process of sodium hydroxide cooking, used in pulp and paper industry. Additionally we altered the chemical and physical properties of those substrates by drying and rewetting, treatment of sodium hydroxide and sodium chlorite in order to decrease or increase the surface area and delignify, respectively. We measured cellulolytic enzyme activity by enzymatic assays. Proteins were examined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional gel electrophoresis. The enzymes produced were subsequently used for enzymatic hydrolysis of lignocellulosic substrates and compared to enzymatic hydrolysis of model cellulosic substrates, namely, Avicel PH101, nanocrystaline cellulose, phosphoric acid-swollen cellulose and cotton, which have defined characteristics. The structural properties of the substrates during the different times of hydrolysis were analyzed by solid-state nuclear magnetic resonance (NMR) technique. Dynamics of the hydrolysis was analyzed by quartz crystal microbalance with dissipation (QCM-D) technique. Hydrolysis products were verified by high performance anion-exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD)