An investigation on the performance of hydrogen production by simultaneous saccharification and fermentation (SSF) in a dual-chamber microbial electrolysis cell (MEC) was carried out to consider different anolyte pH levels and culture temperatures, and the influences of anolyte pH value and culture temperature on changes of current, organic acid and pH value were also evaluated. The maximal hydrogen production rate (HPR) of 2.46 mmol/L/D (hydrogen energy recovery 219.02%) was obtained at the initial anolyte pH of 6.5. Within the range of the tested operation temperatures (30-50 degrees C), the optimal temperature for hydrogen production by SSF in the MEC systems was 35 degrees C. Moreover, the contents of organic acids and reducing sugar significantly changed with varying in initial anolyte pH and temperature levels. The result indicates that a low initial anolyte pH value and high culture temperature was beneficial to hydrolysis of cellulose, and a high initial anolyte pH value and a moderate culture temperature to hydrogen production. (C) 2017 Hydrogen Energy Publications LLC. Published by Elsevier Ltd. All rights reserved.