To improve the hydrogen evolution rate in continuous hydrogen production of a novel fermen-tative hydrogen-producing bacteria st rain B49 (AF481148 in EMBL), 4% immobilized cells by polyvinyl alcohol-boric acid method, with the addition of a small amount of calcium alginate in a column reactor obtain hydrogen yield of 2.31mol H2/mol glucose and hydrogen evolution rate of 1435.4ml/L culture•h respectively at medium retention time of 2.0h with a medium containing 10g glucose/L. Moreover, as the cell density in gel beads is increased to 8%, hydrogen yield and hydrogen evolution rate for 10g glucose/L are 2.34mol H2/mol glucose and 2912.4ml/L culture•h respectively at medium retention time of 1.0h, and for molasses wastewater COD of 7505.9mg/ L hydrogen production potential of 205.6ml/g COD and hydrogen evolution rate of 2057.7ml/L culture•h at hydraulic retention time of 0.75h are observed. In the continuous culture pH value keeps around 3.9 by self-regulating.