NH(4)(+), which is normally the integrant in organic wastewater, such as Tofu wastewater, is an inhibitor to hydrogen production by anoxygenic phototrophic bacterium. In order to release inhibition of NH(4)(+) to biohydrogen generation by Rhodobacter sphaeroides, a glutamine auxotrophic mutant R. sphaeroides TJ-0803 was obtained by mutagenizing with ethyl methane sulfonate. The mutant could generate biohydrogen efficiently in the medium with high NH(4)(+) concentration, because the inhibition of NH(4)(+) to nitrogenase was released. Under suitable conditions, TJ-0803 could effectively produce biohydrogen from tofu wastewater, which commonly containing 50-60 mg L(-1) NH(4)(+), and the generation rate was increased by more than 100% compared with that from wild-type R. sphaeroides. (C) 2010 Elsevier Ltd. All rights reserved.