Following pretreatment and enzymatic hydrolysis of lignocellulosic biomass within a biorefinery, the residual solids are often present in high concentration and have a wide particle size distribution. These solids, which can be difficult to remove from solution, can have a detrimental impact on downstream fermentations and separations. Here we show that polyelectrolyte flocculating agents can be used to enhance the solid-liquid clarification of a lignocellulosic biomass (Ponderosa Pine) hydrolyzate. Due to the negative zeta potential of Ponderosa Pine solids following pretreatment and enzymatic hydrolysis, flocculation with cationic polymers provided the largest flocs, which quickly settled out of solution. Adding Kemira flocculant C1592 up to 1000 mg/L created larger flocs; however, a higher dosage (5000 mg/L) resulted in solids re-dispersing into solution. It was estimated that a greater than 12-fold improvement in throughput with a scroll decanting centrifuge could be obtained when using flocculant, or that an approximately 40-fold higher flux could be obtained for a vacuum filtration operation. The addition of 100 mg/L of C1592 showed optimal suspended solids removal of the clarified solution (<0.1% w/w suspended solids) and the highest sugar recovery (up to 83% without washing). In all cases, the percentage of suspended solids remaining following either centrifugation or filtration was at least an order of magnitude lower with the aid of flocculant. The presence of residual polymer in solution did not affect the downstream ethanol productivity of a Saccharomyces cerevisiae fermentation. (C) 2010 Elsevier Ltd. All rights reserved.