Research On Bio-Hydrogen Production By Two-Step Incubation Of Dark-Fermentative Bacterium And Photo-Fermentative Bacterium

As advanced renewable energy, hydrogen has been received more and more concerns of the whole world. Among most methods of producing hydrogen, using microbe to ferment biomass such as organic waste water, agriculture solid wastes and organic rubbish in anaerobic condition to produce hydrogen is becoming a noticeable problem all over the world, because of the advantage of getting rid of wastes, protecting environment and producing energy. And an integrated biological H2 production of the dark-fermentative bacterium and the pH oto-fermentative bacterium has become a subject of considerable interest.Since the pHoto-fermentative bacterium can utilize the volatile fatty acids (VFAs) produced by the dark-fermentative bacteria, this process can release the inhibition of VFAs in the dark-fermentation, increase the substrate utilization and realize continuous and stable H2 production. However, this process of integrated biological H2 production has not been investigated extensively and stayed in the stage of library. And the low efficiency of substrate utilization and difficulties in co-culture cannot be ignored. Basing on the standpoint of interaction in biohydrogen production, we screened out strains of pH oto-fermentative bacterium that could produce hydrogen with high efficiency, and mixed them with Clostridium butyricum, hoping to achieve highly hydrogen producing capability, and discuss the key factors that influence the interaction between the mixed bacterium.We isolated a strain pH oto-fermentative bacterium (strain RLD119) that can produce hydrogen. Through the test of 16S rDNA, the most similar bacterium with strain RLD119 is Rhodopseudomonas palustris, and the comparability is about 99%.Though the hydrogen producing ability comparison of the three pH oto-fermentative bacteria Rhodopseudomonas palustris, Rhodobacter capsulate and Rhodopseudomonas.sp utilizing different substrates, all of them could utilize the VFAs and glucose to grow, but only Rhodopseudomonas.sp can produce hydrogen which was instable. On the other hand, strain RLD119 can utilize various substrates to grow, and acetate, propionate, and glucose to produce hydrogen, with the highest hydrogen yield utilizing acetate. In the mixed substrates of acetate and butyrate with the proportion of 1:1, the highest hydrogen yield with 1916 ml/L was obtained.In the two-step culture of Clostridium butyricum and RLD119, the manner of which the culture broth of Clostridium butyricum was regulated with pH and autoclavd, has the highest H2 evolution, in yields of 5.5 mol H2/mol glucose, with the control of 1.8 mol H2/mol glucose of the Clostridium butyricum, and 1.1 mol H2/mol glucose of RLD119. When they were co-cultured with different proportion, the 1: 600 mixed culture between Clostridium butyricum and RLD-119 has the highest H2 evolution, in yields of 1.97 mol H2/mol glucose.Therefore, the two-step culture with regulated pH and autoclaving is the most efficient manner of integrated biological H2 production The influence of inoculating proportion and light intensity on hydrogen producing ability in two-step culture was reviewed. Under the inoculating proportion of Clostridium butyricum and RLD-119 between 5:2 and 5:3, the hydrogen producing efficiency was far better than that of the other proportion, and the highest hydrogen yield with 1560ml/L was obtained at the inoculating proportion of 5:3. Under the condition of 7500lx of light intensity, the best result was achieved with 2640ml/L and 4.9molH2/mol glucose.