Pretreatment is being the first and most expensive step, it has pervasive impacts on all other steps in overall conversion process. There are several pretreatment methods using physical, chemical, and biological principles which are under various stages of investigation. Extrusion can be used as one of the physical pretreatment methods towards biofuel production. The objective of this study was to evaluate the effect of barrel temperature and screw speed on sugar recovery from corn stover, to select a suitable enzyme combination and its ratio. Corn stover was pretreated in a single screw extruder with five screw speeds (25, 50, 75, 100, and 125 rpm) and five barrel temperatures (25, 50, 75, 100, and 125 degrees C). In order to select a suitable enzyme combination and ratio, different levels of cellulase and beta-glucosidase, multienzyme complex and beta-glucosidase were used during saccharification of pretreated corn stover. From the statistical analysis, it was found that screw speed and temperature had a significant effect on sugar recovery from corn stover. Higher glucose, xylose, and combined sugar recovery of 75, 49, and 61%, respectively, were recorded at 75 rpm and 125 degrees C. This pretreatment condition resulted in 2.0, 1.7, and 2.0 times higher than the control sample using 1:4 cellulase and beta-glucosidase combination.