Optimal conditions for the cold alkaline extraction (CAE) pretreatment of eucalyptus, sugarcane bagasse and sugarcane straw are proposed in view of their subsequent bioconversion into ethanol through the semi -simultaneous saccharification and fermentation (SSSF) process (with presaccharification followed by simultaneous saccharification and fermentation, or SSF). The optimum conditions, which are identified based on an experiment with a factorial central composite design, resulted in the removal of 46%, 52% and 61% of the xylan and 15%, 37% and 45% of the lignin for eucalyptus, bagasse and straw, respectively. The formation of pseudo-extractives was observed during the CAE of eucalyptus. Despite the similar glucose concentration and yield for all biomasses after 12 h of presaccharification, the highest yield (0.065 g(ethanol)/g(biomass)), concentrations (5.74 g L-1) and volumetric productivity for ethanol (0.57 g L-1 h(-1)) were observed for the sugarcane straw. This finding was most likely related to the improved accessibility of cellulose that resulted from the removal of the largest amount of xylan and lignin. (C) 2016 Elsevier Ltd. All rights reserved.