Caldicellulosiruptor saccharolyticus is an extreme thermophilic bacterium recognized for its saccharolytic ability and superior ability to produce high yields of hydrogen. However, most studies have been made using yeast extract (YE) as a rich but expensive nutrient source. For the first time, we show that C. saccharolyticus is able to grow on defined minimal media, including essential vitamins, provided that CO2 was allowed to accumulate sufficiently in the culture broth to activate growth. Growth and hydrogen production performance on minimal media was analyzed in both batch and continuous mode. Absence of YE resulted in similar or higher hydrogen yields and specific hydrogen productivities but lower volumetric hydrogen productivities than with YE. The results also indicate that YE is used as a carbon- and energy source thus affecting metabolic flux calculations. This study clarified that YE is not essential making C. saccharolyticus more attractive for fundamental studies on its metabolism and future industrial exploitation. Copyright (C) 2011, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.