This study demonstrates that, besides sulfur deprivation, sustained H-2 photoproduction in Chlamydomonas reinhardtii cultures can be generated by incubating algae under phosphorus-deprived (-P) conditions. However, phosphorus deficiency in algal cells could not be obtained by resuspension of algae in -P medium, evidently due to a significant reserve of phosphorus in cells. In this study, phosphorus deficiency was accomplished by inoculating the washed algae into the -P medium at low initial cell densities (below 2 mg Chl l(-1)). After the initial growth period, where cells utilize intracellular phosphorus, algae established anaerobic environment followed by the period of H-2 photoproduction. The maximum H-2 output (similar to 70 ml l(-1)) was obtained in cultures with the initial Chl content similar to 1 mg l(-1). Cultures with Chl above 2 mg l(-1) did not produce H-2 gas. The physiological response of algal cultures to phosphorus deprivation demonstrated significant similarities with the response of algae to sulfur depletion. Copyright (C) 2012, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.