2.5. Enzymatic Hydrolysis Test

The WIS obtained after extrusion of VTPW was enzymatically hydrolyzed using a commercial enzyme blend (Cellic Ctec2) purchased from SIGMA (Ref. SAE0020). This preparation contains cellulases, hemicellulases, and β-glucosidase enzymes. Enzymatic hydrolysis assays were performed in triplicate, using 100-mL Erlenmeyer flasks. Assays were run in 50 mM sodium citrate buffer (pH 5) at 5 and 10% (w/v) substrate concentration (dry weight, DW), 50 °C, 150 rpm, and enzyme loading of 15 FPU/g DW of substrate. Representative samples were taken after 24, 48, and 72 h and centrifuged at 9300× g for 10 min. Supernatants were analyzed by HPLC to determine the sugars concentration.

Enzymatic hydrolysis yields (EH) were estimated considering the glucose/xylose produced during enzymatic hydrolysis (after subtracting the sugar content from the enzyme preparation), which is referred to the potential glucose/xylose (calculated based on the glucan/xylan content in the WIS) and was expressed as a percentage (denoted EH_G and EH_X). Average values ± SD of the three replicates were presented.

SF-CT was also subjected to enzymatic hydrolysis in 100-mL Erlenmeyer flasks at 10% (w/v) substrate loadings, 50 °C, and 150 rpm for 48 h. With the aim of maximizing enzymatic hydrolysis yields from SF-CT, different enzyme preparations with different hydrolytic activities were used as follows: (1) 1% (w/w) N22086/g DW of substrate; (2) 0.1% NS22119/g DW of substrate; (3) 15 FPU Cellic CTec2/g DW of substrate. NS22086 and NS22119 were both provided by Novozymes (Denmark) and are enzymatic preparations with cellulases, xylanases, and carbohydrases (arabinase, β-glucanase, cellulase, hemicellulase, pectinase, and xylanase) activities, respectively.