One of the main obstacles to the development of advanced forest biorefineries is the optimal fractionation of the plant biomass. The aim of this work was to design a two-stage process for the fractionation of slash pine sawdust: alkaline deresination for extractives removal and sulfuric acid-catalyzed steam explosion (SE) for hemicellulose extraction. SE was carried out in a high-pressure steam reactor under different conditions of temperature (180 degrees C-200 degrees C), time (5 min-10 min) and acid concentration (1: 100 H2SO4 - 3: 100 H2SO4 (on mass of dry wood)) according to a factorial design. Extractives, lignin, carbohydrates, and degradation by-products were quantified. The effect of pretreatment on enzymatic hydrolysis (EH) was also evaluated. SE was very efficient for hemicelluloses extraction (90% at 200 degrees C with 3: 100 H2SO4 (on mass of dry wood) for 5 min) with a high glucan recovery. EH yield increased 6-fold after SE. There was a trade-off between maximizing carbohydrate recovery, hemicellulose extraction, and cellulose digestibility.