Ipomoea aquatica, a tropical plant was used as a new substrate, and the digested sludge (DS) was used as inoculum for biohydrogen production. In order to inhibit the hydrogen consuming bacteria (HCB), the DS was subjected to thermal and acid pretreatment to identify the optimum method. The results showed that thermal pretreatment was better than acid pretreatment. To further investigate the best thermal pretreatment condition of DS, response surface methodology (RSM) was employed. Consecutively, thermal pretreatment at 90 degrees C for 60 min was identified as the optimum pretreatment condition for inoculum. Further, Ipomoea aquatica used as substrate was also optimized under conditions like freezing, boiling, and alkali pretreatment to attain high hydrogen yield (HY). Frozen and dried 1. aquatica demonstrated the highest HY of 217.16 mL/g-VS, which was manifold higher than control and other treatment conditions. The energy consumed in the fermentation process was evaluated which was lesser than energy produced in the process. Furthermore, a practical process was proposed. To the best of our knowledge, it's the first time that I. aquatica was used as substrate to produce hydrogen through an attractive process that could not only benefit the environment by water purification but also contributes to clean energy production. (C) 2016 Elsevier Ltd. All rights reserved.