Even with the improvements noted above, hydrogen yields of fermentative hydrogen production processes are restricted by the existing metabolic pathways to 2 or 4 mol H2/mol glucose consumed, for butyrate or acetate fermentation, respectively. The techniques already discussed in Section 13.3.6 could not increase yields beyond these limits. In practice, typical yields in the range of 1 to 2 mol Hj/ mol of glucose result in 10-20% chemical oxygen demand (COD) removal, since the main part of the organic content of the wastewater remains in the liquid phase in the form of various VFAs and solvents. Even under optimum conditions of 4 mol H2/mol glucose, about 60-70% of the organic matter of the feed remains in solution (Venkata et al., 2008; 2009). Further utilization of the organic matter contained in the effluent of a fermentative hydrogen producing bioreactor, could increase the overall energy output of the process. The development of a two-stage process involves the fermentation of the substrate to hydrogen and organic acids in the first stage and additional energy extraction by feeding the effluent of the first stage reactor to a second stage.

One approach to utilize/reuse the remaining organic matter in producing a second useable form for energy (an energy carrier) is to produce CH4 in a second stage. Integration of an acidogenic process with a subsequent methanogenic process for combined hydrogen and methane generation, offers several advantages such as a higher performance of the process in terms of waste stabilization efficiency and net energy recovery (Ghosh et al., 1985). Such a two-stage system has been proposed so far for organic solid wastes rich in carbohydrates such as food wastes (Han and Shin, 2004), cheese whey (Antonopoulou et al., 2008a; Venetsaneas et al., 2009), olive mill wastewaters (Koutrouli et al., 2009), household solid waste (Liu et al., 2006b), a mixture of pulverized garbage and shredded paper wastes (Ueno et al., 2007a) and wastewater sludge (Ting and Lee, 2007). A combined hydrogen — and methane-generation process has already been scaled up to the pilot plant stage, for organic solid wastes (Ueno et al, 2007b). The hydrogen and methane production rates were 5.4 m3/m3/d and 6.1 m3/m3/d, respectively while the process COD removal efficiency was 80%. The overall efficiency of this combined process is demonstrated by the fact that methane yields were twofold higher than a comparable single-stage process (Ueno et al., 2007b).

Another approach to increase the overall energy extraction is to couple the fermentative hydrogen production with photofermentation with the aim to recover additional hydrogen. In such a two-stage process, the rich in organic acids effluent of fermentation which is produced in the first stage by anaerobic fermentative bacteria could be converted to hydrogen in the second step by non-sulfur purple photosynthetic bacteria which capture light energy, using a photobioreactor. This combination of both kinds of bacteria not only reduces the light energy demand of the photosynthetic bacteria but also enhances the hydrogen yield as well (Das and Veziroglu, 2001). Intensive research has been carried out in this area (Nath et al., 2008; Chen et al., 2008b) in the last few years. However, there are important factors limiting the practical application of such a process. One of them is that the involved hydrogen enzyme, nitrogenase, is potentially sensitive to the nitrogen content of the medium/substrate since nitrogen inhibits enzyme activity, as well as represses nitrogenase synthesis. However, this limitation can be potentially overcome either by genetic manipulation (Drepper et al., 2003) or selection (Rey et al., 2007) to remove nitrogenase regulation. In addition, one of the most severe constraints is that photosynthetic efficiencies are very low since at even moderate light intensities, the main part of captured light is dissipated as heat (Hoekema et al., 2006). This means that there will be a demand for large surface areas for the production of hydrogen contributing to the total cost and render the development of a two-stage process of fermentation-photofermentation, far from practical application.

Another approach to increase the overall energy recovery could be the coupling of fermentation with the additional hydrogen production, via a MEC. In this two — stage system, the organic acids which are typical by-products of hydrogen fermentation will be converted to hydrogen in a MEC (Liu et al., 2005; Rozendal et al., 2006). Specifically, the electrogenic bacteria, catabolize the substrates and use the anodic electrode as terminal electron acceptor while supplementary voltage (>200 mV) is added in order to drive hydrogen evolution at the cathode. Thus, a sequential second stage of a MEC after a fermentative hydrogen production first stage could completely convert the effluent of first step to hydrogen, achieving in principle, 12 mol H2/mol glucose with only a small electricity supply. However, the fact that the yields for MEC which have already been reported in the literature are quite lower than the respective yields of dark anaerobic fermentation process, in combination with the high cost of cathodic electrodes and the reduction of the electrical input, limit the practical applicability of this promising technology.