Biological production of second generation biofuels such as biohydrogen (H-2) or methane (CH4) represents a promising alternative to fossils fuels. Alkaline pretreatments of lignocellulosic biomass are known to enhance the accessibility and the bioconversion of hollocelluloses during anaerobic digestion and dark fermentation processes. In the present study, four different configurations were investigated: one-stage CH4 (continuous) and two-stage H-2 (batch)/CH4 (continuous) process with and without alkaline pretreatment of sunflower stalks (55 degrees C, 24 h, 4 g NaOH/100 g TS). The results showed that two stage H-2/CH4 (150 +/- 3.5 mL CH4 g(-1) VS) did not improve methane yields compared to one stage CH4 (152 +/- 4 mL CH4 g(-1) VS). Although alkaline pretreatment was shown to be inefficient in improving the H-2 yields in the two-stage H-2/CH4 process, an increase in methane yields by 26% and 29% were observed with one-stage CH4 and two-stage H-2/CH4 production compared to one-stage CH4 process without alkaline pretreatment, respectively. Chemical analysis of the solid digestate showed that hemicelluloses were the most preferred substrates compared to cellulose whereas lignin remained undegraded in all four studied configurations. Finally, energy balance showed that a positive energy balance and economic sustainability can be achieved when the alkaline pretreatment is applied at a high substrate concentration and/or when heat is recovered at a maximum efficiency during the pretreatment step. (C) 2014 Elsevier B.V. All rights reserved.