Abstract A combination of alkaline thermal pretreatment followed by thermomechanical extrusion was studied as a novel sequential pretreatment process for an effective breakdown of the lignocellulosic structure of Eucalyptus grandis wood (EW). The first step was studied by analysing the influence of two factors: the NaOH-to-dry biomass ratio or NaOH loading (NaOH/DM) and the liquid-to-solid ratio (L/S). Optimization of these two parameters provided good results in terms of enzymatic hydrolysis at 5% (w w−1) solids loading, obtaining a total sugar concentration of 24.9 g L−1 and a total sugar production of 36.9 g 100 g−1 raw EW after pretreating the biomass at 20% NaOH/DM and L/S = 1/1. The second step of extrusion, when followed by a final washing step, provided a significant increase in glucose and xylose production when working at 10% NaOH/DM. For a soda loading of 20%, there was a clear improvement in sugars conversion yield after extrusion and washing: 71% for glucan conversion and 89% for xylan.