Mixed bacteria were used to improve hydrogen yield from cassava starch in combination of dark and photo fermentation. In dark fermentation, mixed anaerobic bacteria (mainly Clostridium species) were used to produce hydrogen from cassava starch. Substrate concentration, fermentation temperature and pH were optimized as 10.4 g/l, 31 degrees C and 6.3 by response surface methodology (RSM). The maximum hydrogen yield and production rate in dark fermentation were 351 ml H(2)/g starch (2.53 mol H(2)/mol hexose) and 334.8 ml H(2)/l/h, respectively. In photo fermentation, immobilized mixed photosynthetic bacteria (PSB, mainly Rhodopseudomonas palustris species) were used to produce hydrogen from soluble metabolite products (SMP, mainly acetate and butyrate) of dark fermentation. The maximum hydrogen yield in photo fermentation was 489 ml H(2)/g starch (3.54 mol H(2)/mol hexose). The total hydrogen yield was significantly increased from 402 to 840 ml H2/g starch (from 2.91 to 6.07 mol H(2)/mol hexose) by mixed bacteria and cell immobilization in combination of dark and photo fermentation. (C) 2010 Professor T. Nejat Veziroglu. Published by Elsevier Ltd. All rights reserved.