Dark fermentative hydrogen production with crude glycerol from biodiesel industry using indigenous hydrogen-producing bacteria Yung-Chung Lo a,d , Xue-Jiao Chen a,b , Chi-Yu Huang c , Ying-Jin Yuan b , Jo-Shu Chang a,d,e, * a Department of Chemical Engineering, National Cheng Kung University, Tainan, Taiwan b School of Chemical Engineering and Technology, Tianjin University, Tianjin, China c Department of Environmental Science and Engineering, Tunghai University, Taichung, Taiwan d Center for Biosciences and Biotechnology, National Cheng Kung University, Tainan, Taiwan e Energy Technology and Strategy Center, National Cheng Kung University, Tainan, Taiwan article info Article history: Received 31 March 2013 Received in revised form 13 May 2013 Accepted 15 May 2013 Available online xxx Keywords: Biohydrogen Glycerol Biodiesel Clostridium butyricum Clostridium pasteurianum Klebsiella sp. abstract Glycerol is an inevitable by-product from biodiesel synthesis process and could be a promising feedstock for fermentative hydrogen production. In this study, the feasibility of using crude glycerol from biodiesel industry for biohydrogen production was evaluated using seven isolated hydrogen-producing bacterial strains (Clostridium butyricum, Clos- tridium pasteurianum, and Klebsiella sp.). Among the strains examined, C. pasteurianum CH4 exhibited the best biohydrogen-producing performance under the optimal conditions of: temperature, 35  C; initial pH, 7.0; agitation rate, 200 rpm; glycerol concentration, 10 g/l. When using pure glycerol as carbon source for continuous hydrogen fermentation, the average H 2 production rate and H 2 yield were 103.1  8.1 ml/h/l and 0.50  0.02 mol H 2 /mol glycerol, respectively. In contrast, when using crude glycerol as the carbon source, the H 2 production rate and H 2 yield was improved to 166.0  8.7 ml/h/l and 0.77  0.05 mol H 2 /mol glycerol, respectively. This work demonstrated the high potential of using biodiesel by- product, glycerol, for cost-effective biohydrogen production. Copyright ª 2013, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved. 1. Introduction The combustion of fossil fuels gives serious negative effects on the environment mainly because of the emissions of green- house gases (e.g., CO, CO 2 ) and air pollutants (e.g., NOx, SOx and fly ash). For these reasons, many researchers have been searching new sustainable energy sources that could replace fossil fuels [1]. Biomass energy is a promising alternative to fossil fuels. It is produced by microbial conversion of biomass, such as agricultural, forestry, husbandry wastes or lignocel- lulosic materials to methane, hydrogen, ethanol, gasoline or biodiesel. In the past decades, hydrogen, the intermediate of anaerobic digestion process, has gained increasing attention since it is clean, efficient, and sustainable [2e4]. Biological hydrogen production is considered the most environmentally friendly way of producing energy [5e13]. * Corresponding author. Department of Chemical Engineering, National Cheng Kung University, No. 1 University Road, Tainan 701, Taiwan. Fax: þ886 6 2357146, þ886 6 2344496. E-mail address: changjs@mail.ncku.edu.tw (J.-S. Chang). Available online at www.sciencedirect.com journal homepage: www.elsevier.com/locate/he international journal of hydrogen energy xxx (2013) 1 e8 Please cite this article in press as: Lo Y-C, et al., Dark fermentative hydrogen production with crude glycerol from biodiesel industry using indigenous hydrogen-producing bacteria, International Journal of Hydrogen Energy (2013), http://dx.doi.org/ 10.1016/j.ijhydene.2013.05.083 0360-3199/$ e see front matter Copyright ª 2013, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.ijhydene.2013.05.083