Chlamydomonas reinhardtii UTEX 90 accumulated 1.45 g dry cell weight and 0.77 g starch/L during photosynthetic growth using TAP media at 25 degrees C in presence of 2% CO2 for 3 days. C. reinhardtii biomass was concentrated and then converted into hydrogen and organic acids by anaerobic fermentation with Clostridium butyricum. Organic acids in the fermentate of algal biomass were consecutively photo-dissimilated to hydrogen by Rhodobacter sphaeroides KD131. In the concentrated algal biomass 52% of the starch was hydrolyzed to 37.1 mmol H-2/L-concentrated algal biomass and 13.6, 25.5, 7.4 and 493 mM of formate, acetate, propionate, and butyrate, respectively by C. butyricum. R. sphaeroides KD131 evolved 5.72 mmol H-2 per ml-fermentate of algal biomass under illumination of 8 klux at 30 degrees C. Only 80% of the organic acids, mainly butyrate, were hydrolyzed during photo-incubation. During anaerobic conversion, 2.58 mol H-2/mol starch-glucose was evolved using C. butyricum and then 5.72 mol H-2/L-anaerobic fermentate was produced by R. sphaeroides KD131. Thus, the two-step conversion process produced 8.30 mol H-2 from 1 mol starch-glucose equivalent algal biomass via organic acids. 0 2005 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.