Biodiesel manufacture produces crude glycerol as a major byproduct. At the scale estimated for future biodiesel production, extensive quantities of crude glycerol fraction will be generated, creating a large waste stream with potentially significant environmental impacts. The magnitude of projected future crude glycerol supplies suggests that its conversion to a biofuel is the only viable route to producing a product that does not cause market saturation. Previously it was shown that crude glycerol could be converted to hydrogen, a possible future clean energy carrier, by photofermentation using Rhodopseudomonas palustris through photofermentation. Here, the effects of nitrogen source and different concentrations of crude glycerol on this process were assessed. At 20 mM glycerol, 4 mM glutamate, 6.1 mol hydrogen/mole of crude glycerol were obtained under optimal conditions, a yield of 87% of the theoretical, and significantly higher than what was achieved previously. Copyright (C) 2011, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.