For global environmental considerations, production of hydrogen by biological reactions from renewable organic waste sources represents an important area of bioenergy production. Many investigations have been conducted using pure cultures of hydrogen-producing bacteria. Some of the studies showed promising results. However, since there is bacterial contamination in wastewater, it is difficult to maintain pure cultures in a wastewater treatment reactor. Therefore the treatment of wastewater usually has to be a mixed cultures process. Although some researchers have tried using mixed cultures, none of their experiments attained stable continuous hydrogen production - the hydrogen production was stopped by either microbial shift or metabolic alteration. So the present study was aimed at developing a feasible anaerobic hydrogen fermentation process, in which the consistency of the enriched mixed culture can be maintained. Environmental conditions such as pH, feeding substrate and intermediates production were also evaluated to maximize hydrogen production. In this study, both continuous flow experiments and batch tests were employed. Non-sterile sucrose/starch solution was used as substrate for the experimental study. Through a 5-stage operation of the continuous flow reactors, it was found that with the addition of a sludge activation and return process, a consistent hydrogen production could be attained. The maximum hydrogen production obtained for the continuous flow reactors was 6.5 L/day for the reactor with an organic loading rate of 10.8 g COD/L-day and 3.5L/day for the reactor with an organic loading rate of 7.1 g COD/L-day. A high loading was found to be more favorable for hydrogen production. Under the same operation conditions, the hydrogen conversion potential from sucrose was higher than that from starch. The pH had a significant effect on both hydrogen production potential and hydrogen production rate. The optimum pH range for hydrogen production obtained from batch tests was between 5 and 5.5. Major intermediates such as propionate, acetate and butyrate detected in the process could also affect hydrogen production.