Liquid AFEX Pretreatment and Enzymatic Hydrolysis of Switchgrass from Different Harvest and Storage Conditions

Abstract. Switchgrass harvest and storage conditions increasingly affected a representative biochemical process and theoretical ethanol yield. The approach used a novel liquid ammonia fiber explosion (LAFEX) pretreatment technique implemented through engineered introduction of ammonia in liquid phase to biomass with controlled pressures, temperatures, and a mechanical liquid pump. The LAFEX pretreatment involved pressurizing an AFEX reactor with nitrogen prior to adding ammonia, and then nitrogen pressure was increased as the reactor temperature was increased to 100°C after the addition of ammonia. LAFEX pretreatment improved Alamo switchgrass xylan digestibility up to 63.3% compared to AFEX pretreatment that had published levels of xylan digestibility up to 38.3%, all using Accellerase enzyme for enzymatic hydrolysis. This improvement occurred at less water loading (0.8 vs. 2.0 g H 2 O per g dry switchgrass), less ammonia loading (1.0 vs. 1.6 and 2.0 g NH 3 per g dry switchgrass), lower temperature (100°C vs. 160°C and 150°C), and increased pretreatment time (120 vs. 30 and 25 min) for LAFEX vs. AFEX pretreatments, respectively. Additional potential for LAFEX pretreatment was noted based on published studies documenting enzymes that outperform the enzyme selected for this study. A wide range of switchgrass field harvest conditions, including one-cut versus two-cut strategies and harvest times ranging from June to January, indicated relatively small differences in fresh switchgrass cellulose and hemicellullose levels (64% to 68%) determined with wet chemistry, and associated trends in lignin increases (21.9% to 25.7%) and ash decreases (5.3% to 2.2%) across samples obtained at harvest time, not including storage effects. Baseline theoretical ethanol yield for freshly harvested samples ranged from 462 to 485 L per dry Mg. Field storage conditions included covered and uncovered bales and piles of field-chopped switchgrass and ensiled field-chopped switchgrass. Field storage conditions affected cellulose and hemicellulose levels, as determined by wet chemistry, for theoretical ethanol yield (243 to 477 L per dry Mg) to a greater degree than variation due to field harvest conditions. Implementation of enzymatic hydrolysis for 72 h with Accellerase 1000 introduced to LAFEX-pretreated samples represented a biorefinery process for theoretical ethanol yield based on fresh harvest conditions (232 to 270 L per dry Mg) and revealed an exaggerated effect of storage conditions on theoretical ethanol yield (79 to 283 L per dry Mg). Essentially, switchgrass that got wet and stayed wet, compared to switchgrass that stayed dry or was allowed a drying cycle if exposed to precipitation during storage, reduced available cellulose and hemicelluloses (to ~50%) and further reduced the available sugars from enzymatic hydrolysis (to ~30%). It should be noted that no amount of improved pretreatment or enzymatic hydrolysis can overcome or recover the degraded sugars that occurred at the bale bottom under wet conditions. Overall results demonstrated the supply logistics challenge to deliver biomass feedstock with predictable, uniform conditions year-round to allow consistent biochemical conversion at commercial scale. Harvesting and storage conditions could also affect biochemical processes associated with ethanol fermentation that were not investigated.