Intensification Of Cellulase Hydrolysis And Recycling At High-solid Loading

To intensify the hydrolysis efficiency and reduce the production cost of bioethanol during lignocellulose bioconversion, corncob was chosen as the raw biomass to investigate the bioconversion of lignocellulose to ethanol at high solids loading. We focused on solving the problem of low hydrolysis efficiency and high enzyme usage, especially at high solids loading, including the development of the intensification approaches of enzymatic hydrolysis and the design of high performance cellulase recycling. 1. Intensification of surfactants on enzymatic hydrolysis of cellulose: Various surfactants were chosen to study their effects on enzymatic hydrolysis of both Avicel and corncob, showing that the hydrolysis efficiencies can be enhanced by non-ionic surfactant. Glucose yield from corncob treated by H2SO4-SAA pretreatment increased to 78.5% with the optimized Tween 80 concentration of 0.1% at cellulase loading of 10 FPU/g cellulose, which was higher than that(65.0%) without Tween 80 at 30 FPU/g cellulose. The results indicated that non-ionic surfactant could effectively increase the efficiencies of enzymatic hydrolysis and thus decrease enzyme usage. The adsorption kinetic of protein proved that cellulase adsorption on non-cellulosic substrate was weakened by Tween 80, whereas it has little effect on cellulase adsorption on cellulose. 2. Design of step-reduction in agitation rate: At various solid loadings, the effect of agitation rate on hydrolysis efficiency of corncob was studied. The obtained results showed that extrame high agitation rate caused cellulase deactivation due to shear force, and the needed agitation rates would be decreased at lower solid loadings. Based on the relationship of agitation rate and hydrolysis efficiency at diffenent solid loadings, we proposed the mixing mode of stepwise reduction of agitation rate at high solid loading, which could obtain high glucose yield and reduce energy consumption for mechanical mixing. 3. Reducing β-glucosidase supplementation in enzyme recycling process using engineered yeast: An engineered yeast expressing β-glucosidase was used for cellulosic ethanol production, which could reduce the amount of supplemented β-glucosidase in enzyme recycling with re-adsorption. SSF of pretreated corncob was conducted using engineered strain at cellulase loading of 30 FPU/g cellulose with dry matter of 10%. Afterwards, cellulase was recovered with the addition of 20CBU/g cellullose β-glucosidase by re-adsorption, the first round cellulase recovery was 92.2%. The amount of β-glucosidase supplementation was reduced to one third of previous supplement. 4. Optimation of cellulase recycling strategy using engineered strain and surfactant: The optimization of cellulase recycling efficiencies was obtained, when the cellulase in liquid phase was recovered by re-adsorption and the bound cellulase to residue was directly reused by recycling insoluble solid. The combination of the optimized cellulase recycling strategy and engineered strain was employed to increase enzyme recycle efficiency and reduce β-glucosidase supplementation. The addition of Tween 80 further enhanced the enzyme recycle efficiency. The cellulase was recovered after SSF of H2SO4-SAA treated corncob at 30 FPU/g cellulose with dry matter of 10%.The second round of cellulase recovery was 88.8% without β-glucosidase supplementration by above enzyme recycling strategy using engineered strain in the presence of 0.1% Tween80.