Hydrogen Production Characters And Hydrogen Metabolism Of Clostridium Beijerinckii RZF-1108

Fermentative biological hydrogen production using organic wastewater and biomass promises to be an economical and sustainable technology for energy source generation, and becomes one of the research hotspot in the field of environment and energy in recent years. Hydrogen-producing bacteria is one of the most important components in fermentative bio-hydrogen process. Isolating and screening high efficiency hydrogen-producing bacteria, optimizing culture conditions and analyzing functional gene expression, understanding the physiological and ecological characteristics of hydrogen-producing bacteria, improving the hydrogen production rate and substrate conversion rate, and genetic transformation will be favorable for enhancing hydrogen production efficiency and reducing cost of bio-hydrogen in large-scale.In this study, 11 hydrogen-production strains were isolated from bio-hydrogen reactor,with butyrate-type fermentation using the modified Hungate roll-tube technique. The strain named RZF-1108 had higher hydrogen production ability among all isolates and had been further studied the 16S rRNA sequence of strain RZF-1108 had 99% similarity with closest relative Clostridium beijerinckii NCIMB 8052. The physiological and morphological characteristic also proved that the strain RZF-1108 belonged to C. beijerinckii species. Thus, it was characterized as a new strain and designated as Clostridium beijerinckii strain RZF1108.The optimum culture condition was 8% of inoculant volume, initial pH 7.0, 10 g/l of glucose concentrations, 0.1 g/l of Mg2+ concentration (MgCl2·6H2O), 0.2 g/l of Fe2+ concentration (FeSO4·7H2O). In batch culture within 24 h, the average production rate of 86 ml H2/g-drycell·h, the maximum hydrogen yield of 1.96 mol H2/mol glucose were obtained.The effect of peptone, beef extract and yeast extract as the sole nitrogen source and various combinations on hydrogen production by strain RZF-1108 was investigated. The results indicated that nitrogen source had a substantial effect on hydrogen production by C. beijerinckii RZF-1108. The single nitrogen source tests indicated that 4 g/l of peptone, 4 g/l of beef extra and 3 g/l of yeast powder are the optimal concentrations, respectively. When yeast extract concentration was 3 g/l, the maximum H2 yield of 1.76 mol H2/mol glucose was obtained. The hydrogen production ability of strain RZF-1108 using mixed nitrogen source was higher than that obtained using single one, the maximum hydrogen yield is 1.85 mol H2/mol glucose, and the average hydrogen production rate is 74 ml H2 / g-drycell·h when using 3 g/l of yeast extract and 2 g/l of beef extract.Quantitative reverse transcriptase PCR (qRT-PCR) analyses showed that different nitrogen sources and their concentrations affected hydA gene expression of RZF-1108. The maximum hydA gene expression is 8×1010 when using 3 g/l of yeast extract and 2 g/l of beef extract, and it was three times higher than that obtained under low hydrogen production when using 3 g/l of peptone and 2 g/l of beef extract. The resluts indicated that hydA gene expression was obvious correlation with hydrogen production, implying that high hydrogen production ability resulted from high expression of hydA gene.