The goal of this study was to sequential fermentation of hydrogen and methane from sugarcane bagasse (SCB). Steam explosion conditions for pretreating SCB were optimum at 195 degrees C and 1.5 min, which yielded 36.35 g/L of total sugar and 2.35 g/L of total inhibitors. Under these conditions (all in g/L): glucose, 11.33; xylose, 24.41; arabinose, 0.61; acetic acid, 2.33; and furfural, 0.02 were obtained. The resulting hydrolysate was used to produce hydrogen by anaerobic mixed cultures. A maximum hydrogen production rate of 396.50 mL H-2/L day was achieved at an initial pH of 6 and an initial total sugar concentration of 10 g/L. The effluent from the hydrogen fermentation process was further used to produce methane. Response surface methodology with central composite design was used to obtain the suitable conditions for maximizing methane production rate (MPR). An MPR of 185.73 mL/L day was achieved at initial pH, Ni and Fe concentrations of 7.59, 3.61 mg/L and 8.44 mg/L, respectively. Total energy of 304.11 kJ/L-substrate was obtained from a sequential fermentation of hydrogen and methane. This approach will not only add value to SCB, in the form of safe and clean energy, but also provide a solution for making use of this abundant waste. (C) 2018 Hydrogen Energy Publications LLC. Published by Elsevier Ltd. All rights reserved.