The effects of hydraulic retention time (HRT) on biohydrogen production and its mixed anaerobic microbial community grown with glucose were investigated in a continuous stirred tank reactor culture. Starting from a HRT of 50 h the culture was acclimated by stepwise HRT reduction until a steady-state was reached at a HRT of 8 h after 19 days. After that the culture was run with HRTs of 6 h, 8 h, 10 h and 12 h, each lasting 9-15 days. Hydrogen, CO2 and dissolved products were daily measured. The species composition of the culture was ascertained using 16S rDNA genes separated by denaturing gradient gel electrophoresis (DGGE). While a hydrogen yield of 1.6 mol/mol glucose was found during the first steady-state at 8 h HRT it stabilized at 1.9 in the following steady-states. The predominant dissolved products were butyrate and acetate in a ratio of about 2.1: 1 and amounted to 82-94% of the total products. In the first 8 h HRT period also propionate was found in an amount of 9%. It is concluded that the increase of the hydrogen yield after transition from 8 h to 6 h HRT is caused by a washout of the propionate producing population during the 6 h HRT period. In fact the fading of two 16S rDNA gene fragments was noticed in the DGGE profiles at the same time. (c) 2006 Elsevier Ltd. All rights reserved.