A 3-L laboratory scale hydrogen fermentor with fill-and-draw operation defined as intermittent-continuous stirred tank reactor (I-CSTR) was established. There were four operational periods included in this study. In run 1 during the acclimation period, corn starch was added as the auxiliary substrate. The feedstocks of the next two periods were corn starch and kitchen mixture, the Taipei kitchen waste and the Kaohsiung kitchen waste, respectively. Compared between these two periods, run 3 fed with Kaohsiung kitchen waste gives a higher hydrogen producing rate of 27 mmol/L/day than run 2. At run 4 that loading rate was increased and hydraulic retention time was decreased. The maximum hydrogen producing rate of 118 mmol/L/day was found. By the detection of terminal restriction fragment length polymorphism (TRFLP) method, some species of Clostridium such as Clostridium stercorarium, Clostridium thermolacticum, Clostridium aldrichii, Clostridium cellobioparum, Clostridium termitidis (cluster III) and Clostridium formicoaceticum (cluster XI) were presented in all the operational periods. Furthermore, the anaerobic batch test for the substrate utilizing efficiency of the sludge taken from the fermentor was also detected. In the test, the maximum hydrogen producing rate of 48 mmol/L/day was detected. The VSS and the total carbohydrate removal were 35% and 66%, respectively. organic nitrogen had only a little fraction converted to ammonia, but the oil and grease were not degraded. (c) 2007 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.