High solids enzymatic hydrolysis of sugarcane bagasse at a relatively low enzyme loading

Abstract

High cellulase loading is still a major impediment in the production of fermentative sugars from high solids enzymatic hydrolysis of lignocellulosic substrates in the enzyme-based “biorefinery” industry. Therefore, the work described in this thesis focused on high solids enzymatic hydrolysis of sugarcane bagasse at a relatively low enzyme loading. First, to enhance the accessibility and hydrolyzability of cellulose imbedded in the lignocellulosic substrate to the cellulase enzymes, the sugarcane bagasse was pretreated with alkaline-catalyzed atmospheric glycerol organosolv (al-AGO) which was proved to deconstruct the recalcitrant cell wall of lignocellulosic biomass with good component selectivity. Second, to run the high solids enzymatic hydrolysis, some key variables such as cellulase loading and initial solid loading were selected for the enzymatic hydrolysis of al-AGO pretreated substrate. Then, to circumvent the drawbacks derived from the high solids content such as high viscosity and poor mass transfer, high solids enzymatic hydrolysis of substrates was explored with fed-batch mode. Finally, the fed-batch enzymatic hydrolysis was studied with the use of additives and accessory enzymes to efficiently achieve high fermentable sugars. Results showed that the high solids enzymatic hydrolysis of alkaline-catalyzed atmospheric glycerol organosolv (al-AGO) pretreated sugarcane bagasse was initiated with a solids content of 8%. Thereafter, 4% of the additional substrates were consecutively added into the hydrolysis system after 6h, 12h, and 18 h to reach a final solid content of 20%. All the cellulase loading (3 FPU/g substrate), supplemented with optimized additives (Tween 80, bovine serum albumin (BSA) & tea saponin) and accessory enzymes (endo-xylanase and lytic polysaccharide monooxygenases (AA9) was added wholly at the beginning of hydrolysis reaction. Further, the enzymatic hydrolysis of al-AGO pretreated substrate at 30% (w/v) solid loading was also carried out. The results showed that the additive mixture (40 mg Tween 80 + 10 mg tea saponin + 20 mg/ g substrate BSA) contributed towards a 30% increase in cellulose hydrolysis after 48 h at 20% solids content. The combination of these additives and accessory enzymes in the high solids enzymatic hydrolysis system remarkably boosted the sugars release, at 72 h of hydrolysis, the enzymatic hydrolysis at 20% solids content reached 83% with 105 g/L of glucose titre and 51 g/L of xylose content. Furthermore, the enzymatic hydrolysis at 30% solids loading reached 69% with 126 g/L of glucose titre and 56 g/L of xylose content. Our results indicate that the fed-batch process is a favorable model for high solids enzymatic hydrolysis of lignocellulosic substrates. Moreover, the combination of additives and accessory enzymes can greatly boost the high solids enzymatic hydrolysis of lignocellulosic substrates.