# Photo-Fermentative Bacteria Used for Hydrogen Production
## 2 Photo-Fertanative Bacteria for Hydrogen Production
### Rhodopseudomonas
#### 2.2.1 Rhodopseudomonas palustris  
This is a Gram-negative bacterium that uses light energy for its metabolic pathway, which leads to the conversion of organic matter into hydrogen. These microbes can alleviate heavy metal toxicity, improve plant growth, and control various plant pathogens [28]. Several studies have been carried out using these bacteria to analyze hydrogen production during photo-fermentation. In a study conducted by [PERSON] et al., the biochemical kinetics of photo-fermentative hydrogen production were investigated experimentally  
Figure 4: Photo-fermentative hydrogen production in PNSBs. Adapted with permission from Ref. [26]. 2012, [PERSON], [PERSON], [PERSON], [PERSON], [PERSON]—IntechOpen Limited. Figure 3: A general outline of hydrogen production by photo-fermentative bacteria. and numerically in order to optimize the photo-fermetative hydrogen production process. _R. palustris_ showed a maximum specific growth rate of 0.26 h\({}^{-1}\) at an irradiance of 47.4 W/m\({}^{2}\), 30 \({}^{\circ}\)C culture temperature, at 12 h and a pH of 7.0. An initial glucose concentration of 9.9 g/L was used as a substrate in a batch mode study, and gas chromatography was performed to analyze the hydrogen content with the flow rate of argon as carrier gas adjusted to 25 mL/min; a maximum specific hydrogen production rate of 1.63 \(\times\) 10\({}^{-3}\) h\({}^{-1}\) was observed. An increase in the specific hydrogen production rate was obtained with an increment in temperature to 30 \({}^{\circ}\)C, but this decreased to 1.11 \(\times\) 10\({}^{-3}\) h\({}^{-1}\) when the temperature was further increased to 40 \({}^{\circ}\)C, suggesting a reversible inactivation of cellular enzymes and inhibition of hydrogen formation [29]. In a further analysis, mutant strains of _R. palustris_ were synthesized by the manipulation of nitrogen regulatory genes, as it proved to be an effective way of increasing the ammonia tolerance in photosynthetic bacteria to improve hydrogen production. Acetate (20 mM) and butyrate (20 mM) were used as carbon sources for cell growth and hydrogen production at 30 \({}^{\circ}\)C in [PERSON]'s medium with an irradiance of 39.5 W/m\({}^{2}\). The mutant strain, named nifA draT2, was 25 times more tolerant to ammonium than the wild-type strain, demonstrating that manipulation of nitrogen regulatory genes is a reliable way to increase the ammonium tolerance of these bacteria and improve their hydrogen-producing capacity. The hydrogen produced was collected in syringes and analyzed by gas chromatography. The mutant strain nifA draT2 yielded 2744 \(\pm\) 66 mL of hydrogen/L, which was higher than the wild-type strain, indicating that the hydrogen production was enhanced by the nifA mutation [30]. The review by [PERSON] and [PERSON] discussed the immobilization of photosynthetic bacteria, which helped to increase hydrogen production, and different materials, such as agar, glass beads, alginate, and porous glass have been used for the attachment and immobilization of _R. palustris_ in different studies [31]. A strategy to increase the productivity of hydrogen was recently analyzed in _R. palustris_ using lignocellulosic substrate by supplementing it with iron and molybdenum, which acts as a biocatalyst to facilitate electron transfer and ultimately an increase in hydrogen production with a maximum of 2.15 mM/h [32]. A list of hydrogen-producing _R. palustris_ strains in biofilm PBRs is given in Table 1.  
The results of several studies showed that the optimum process conditions for maximizing hydrogen yields from _R. palustris_ strains were close to pH 7 and at a temperature range of 30-40 \({}^{\circ}\)C in different bioreactors and varying light intensities from different sources. Therefore, this microbe has the potential to use different organic sources to produce hydrogen, offering a glimpse into the near future where sustainable and renewable energy sources will play an important role.