A preliminary study of a new route for the valorization of microalgal biomass including protein and bio-fuels recovery is presented. The study involved consecutive steps of soluble protein extraction using alkaline conditions, followed by lipid extraction for biodiesel production. After both extraction processes, the spent biomass was used as feedstock for bio-oil production through pyrolysis at 500 degrees C. The results were compared with pyrolysis using whole microalgal biomass and lignocellulosic feedstock. The new route allowed getting 10% of solubilized protein and only 2% of biodiesel (relative to the total biomass). Bio-oil yield obtained using spent Botryococcus braunii was 33.2%. Higher bio-oil yields were obtained from whole Nannochloropsis gaditana (38.3%), whole B. braunii (39.7%) and pine wood (39.9%). Related to bio-oil characteristics, several protein-derived compounds were identified in bio-oil from spent B. braunii biomass. Therefore, the reduction of these compounds in bio-oil should be a critical target for further research. In addition, considering the low biodiesel production yield, lipid extraction from biomass could be avoided. This would increase lipid derived compounds in bio-oil, improving its heating value. Finally, unlike lignocellulosic biomass derived bio-oil, microalgae bio-oil showed a neutral pH, preventing possible corrosion problems in combustion engines. (C) 2015 Elsevier Ltd. All rights reserved.