High Solids Roller Bottle Enzymatic Hydrolysis and Respirometer Fermentation for High Throughput Screening of Lignocellulosic Biomass

High solids enzymatic hydrolysis increases the final fermentable sugar concentration in the hydrolysate, thereby making the process more economically viable by reducing equipment size and energy requirement. But this method has bottlenecks like improper mixing and low enzyme accessibility, which affects the final sugar concentration, which can be rectified through horizontal mixing on a laboratory scale roller bottle mixer. Ultimately the objective is to make this approach applicable to rapidly identify the effects of abiotic and biotic stresses on different feedstocks under industrially relevant process conditions. The samples were subjected to AFEX (ammonia fiber expansion) pretreatment at 1:1 ammonia loading for corn stover and 2:1 for switchgrass, sorghum, mixed prairie and miscanthus. Enzymatic hydrolysis was carried out on the pretreated samples in 85 mL polycarbonate centrifuge tubes at 6% and 9% glucan loading (approximately 25% and 30% by weight). The biomass was autoclaved prior to enzymatic hydrolysis to limit microbial contamination. An enzyme cocktail of 70% cellulase and 30% hemicellulase was used for these experiments, and the pH was controlled using phosphate buffer. After 72 hours of enzymatic hydrolysis at 50^oC the samples were subjected to centrifugation at 12000 rpm for either 1 or 3 hours. The hydrolysates obtained from these experiments were separately subjected to fermentation by Saccharomyces cerevisiae and Zymomonas mobilis. The hydrolysates and fermentation products were analyzed using HPLC to obtain sugar and ethanol concentrations respectively. Based on the results, 6% glucan loading was found to be more effective considering the final sugar and ethanol yields.