Water hyacinth was treated with microwave-assisted dilute H2SO4 to improve saccharification before enzymatic hydrolysis and H-2 production during dark fermentation. A maximum reducing sugar (RS) yield of 64.4 g/100 g total volatile solid (TVS) (96.1% of the theoretical RS yield) was achieved when water hyacinth was treated through microwave heating with 1% dilute H2SO4 for 15 min at 140 degrees C and then enzymatically hydrolyzed for 72 h. During enzymatic hydrolysis, glucose was efficiently produced from the hydrolysis of cellulose that resulted from the disruption of the lignocellulosic structure of water hyacinth after microwave-assisted H2SO4 treatment. When the hydrolyzed water hyacinth was inoculated with H-2-producing bacteria to produce H-2 during dark fermentation, a maximum H-2 yield of 112.3 ml/g TVS was obtained. The major sugar compositions in the residual solution from dark fermentation were xylose and cellobiose (total RS utilization efficiency: 88.5%). Through a combination of dark fermentation and photofermentation, the maximum H-2 yield from water hyacinth was significantly increased from 112.3 ml/g TVS to 751.5 ml/g TVS, which is 75.2% of the theoretical H-2 yield. (C) 2013 Elsevier Ltd. All rights reserved.