Biofuels from lignocellulosic biomass have the potential to reduce the fossil fuels consumption in the following years. However, the structure of these materials hinders degradation in the hydrolysis stage, which requires pretreatments that helps increase the production of fermentable sugars to obtain biofuel. Therefore, in this study three pretreatments were applied to corn stover biomass: Mechanical, with size reduction (0.5, 1, and 2 mm), followed by an alkaline pretreatment (NaOH 1.5% w/v) and subsequent acid pretreatment at different concentrations (H2SO4 2, 4, and 6% v/v). Finally an acid hydrolysis (1% H2SO4 v/v) of pretreated biomass was applied in order to evaluate the effect of different pretreatments in the amount of total reducing sugars (TRS).