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Pelleting of biomass can increase their bulk density and thus improve storability and reduce transportation costs. The objective of this research was to determine the effects of the pelleting conditions on chemical composition and fermentable sugar yield of the biomass. Corn stover, wheat straw, big bluestem, and sorghum stalks were used for this study. Dilute sulfuric acid was used for biomass pretreatment. Accellerase 1500™ was used for cellulose hydrolysis. Effects of mill screen size, die thickness, and L/D ratio of die on chemical compositions and sugar yield were determined. Glucan content of the biomass was positively affected by die thickness and negatively affected by mill screen size. Opposite trend was observed for xylan content. Wheat straw pellets had the highest sugar yield (92.5-94.1%) and big bluestem pellets had the lowest sugar yield (83.6-91.1%). Optimum pelleting condition is 6.5 mm screen size and 44.5 mm die thickness. [ABSTRACT FROM AUTHOR]