Dried CS (dry weight 1.0 g) was added to the 20 g mixture of
Fenton reagent media containing FeSO4 (0–1 mmol/CS) and H2O2
(8.75–35 mmol/CS) in a 100 mL round bottom flask. To prevent
possible water loss, this flask was sealed with a rubber lid. After
the pretreatment media and CS were first mixed with ratio of
1/20 (w/w) at certain temperature (30–55 C), the magnetic stirring
device was used to ensure uniform temperature in the pretreatment
system. After the pretreatment was over, 20 mL
deionized water as anti-solvent was added to the pretreatment
media. This mixture was stirred vigorously for 10 min, and the
solid regenerated CS was separated by vacuum filtration. Then,
the Fenton solution was separated from the solid residues by filtration
followed by washing the solid CS with deionized water. The
solid regenerated CS were washed repeatedly with deionized
water to remove possible FeSO4, H2O2, and other potential inhibitors
and deactivators of cellulose enzyme hydrolysis (Ximenes
et al., 2010, 2011). Solid CS samples were then dried at 85 C in a
vacuum oven and further used for the enzymatic hydrolysis.