Dark fermentative hydrogen production by Enterobacter aerogenes in a packed bed bioreactor using three immobilization supports was studied. The effects of glucose concentrations (20, 10, 5, 2.5, and 1.5 g/L) and support matrixes (Persian Gulf dead coral, ceramic saddle, and ceramic ball) on the hydrogen yield (HY) and hydrogen production rate (HPR) at low hydraulic retention time (HRT) were investigated. According to the achieved results, glucose concentration, porosity and shape support, and bed voidage affected hydrogen production. The HYs were increased with decreasing glucose concentration and at 2.5 g/L glucose concentration, the maximum HYs of 3.11, 2.86 and 2.79 mol hydrogen/mol glucose were obtained for dead coral, ceramic ball, and ceramic saddle, respectively. The results indicated that at the HRT of 0.6 h, the bioreactor packed with dead coral resulted in higher HY (3.11 mol H2/mol glucose) and HPR (173 mL/L.h) representing 8–11%, and 8–15%, enhancement compared to the bioreactor packed with ceramic supports. Besides, based on the acquired results, the HY and HPR of the ceramic ball were higher than the ceramic saddle.