Characteristics And Hydrogen-Producing Analysis Of Several Purple Non-Sulfur Bacteria

The reduce of finite reserved fossil fuel and the continual increase on energy sources requirement and environment pollution resulted in fossil fuel burning, the problem of energy will be confronted with great challenge in 21st century. So reproducible and clean energy sources will be the principal part of the continuable development energy sources system in the future. Hydrogen is clean and renewable. It only liberates large amount of energy and water after combustion, without evolving the greenhouse gas. Technology with microbe for hydrogen production needs a facile reaction condition and wastage of the reaction system is low. So it gives a solution for the conflict of energy and environment. Photosynthetic-bacteria (PSB) bio-hydrogen production is a green technology of hydrogen production. Culture condition is a very important factor for purple non-sulfur bacteria bio-hydrogen production, because nitrogenase which is one of the key enzymes for Photosynthetic-bacteria bio-hydrogen production is very sensitive to NH3. When there is NH4+ in the environment, the bacteria will not product hydrogen. If the nitrogen source is ammonium, hydrogen output will be reduced; if the nitrogen source is albumin, glutamine, yeast extract and so on, hydrogen output will be enhanced. So optimization of culture condition and picking out the high effective hydrogen production strains are efficiency methods to enhance hydrogen output. In this study, 5 purple non-sulfur bacteria were enriched, screened and identified, based on their physiological and biochemical characteristics; their cultural conditions and the hydrogen-producing ability were determined. According to the cultural morphology, cellular spectra, physiological properties, carbon sources, blast of the sequences of 16S rDNA and analysis of phylogenetic tree, two of the isolated 5 strains were identified as Rhodopseudomonas palustris and other 3 were Rhodobacter sphaeroide. The equations of cell concentration and luminosity were deduced by comparing the quantities of OD600 nm and dry-weight of cell biomass. There are many differences among the 5 strains in utilization abilities of carbon sources, cultural conditions and abilities of hydrogen producing. The most suitable initial pH value, inoculation and yeast concentration were gained and the strain AN2 (Rhodobacter sphaeroide) has the best ability of hydrogen producing. The main contents and parts of conclusion were listed as follow:①Purple non-sulfur bacteria needs one or several vitamins to be growth factor, it is more expensive using vitamin solution than yeast extract , so the culture condition was optimized, using 1‰concentration yeast extract for growth factor to replace the vitamin solution.②Samples were collected from different environment, selective enrichment techniques for phototrophic bacteria were used, RCVBN culture medium was confected to purify isolated strains on the agar plates. 5 strains were purified, named AN1, AN2, AS1, BS1 and D1.③The culture modality of these strains were determined. According to these characters, 5 strains were classified to two genera: AN1 and D1 were been a kind; AN2, AS1 and AS2 were been the other one. Gram staining indicated that all the strains were gram negative bacteria. Pigments are a signify characteristic of the classifying purple non-sulfur bacteria, so the absorption spectrums of live cell suspension of these strains were determined. The carbon and electron sources of 5 strains were set out, all the strains could not growth in the culture medium with 3% NaCl. Based on these characters, AN1 and D1 were identified belonging to Rhodopseudomonas, AN2, AS1 and BS1 belonging to Rhodobacter. Also we had used the 16S rDNA to identify, about 1.4kb sequences were fined in the PCR reaction. After blasting on the NCBI and analysis of the phylogenetic tree, we could make sure that AN1 and D1 was R. Palustris, AN2, AS1 and BS1 was R. sphaeroides.④The equations of cell concentration and luminosity were deduced by comparing the quantities of OD600 nm and dry-weight of cell biomass. The effect of different initial pH value, inoculation and yeast extract concentration for the strains growth were gained, the result showed that D1 grew well when pH value was 5~6, and others needed pH value to be about7.0, the best inoculation was 10% to 20% and it was better that yeast extract concentration was 0.1 to 0.15g/L⑤Hydrogen production experiment indicated the strain AN2 R. sphaeroides has the best ability of hydrogen producing(9.55μg/mL d-1), so we could consider it was a strains with high effective hydrogen production and using it in the after research.