Direct photolysis involves water oxidation and a light-dependent transfer of electrons to the [Fe]-hydrogenase, leading to the photosynthetic hydrogen production. Electrons are derived from water upon the photochemical oxidation by photosystem II (PSII or water — plastoquinone oxidoreductase), which is an enzyme located in the thylakoid membrane of algae and cyanobacteria. PSII uses photons from sunlight to energize electrons that are then transferred through the thylakoid membrane electron-transport chain and, via photosystem I (PSI or ferredoxin oxidoreductase) and ferredoxin (Fd), to the hydrocarbon cluster of [Fe]-hydrogenase (Florin et al., 2001). Plastoquinone is reduced to plastoquinol from the transferred electrons, which are used to reduce NADP+ to NADPH or are used in cyclic photophosphorylation. The energized electrons are replaced by oxidizing water to form hydrogen ions and molecular oxygen, as shown in Figure 9.1. By obtaining these electrons from water, PSII provides the electrons needed for the photosynthesis. The hydrogen ions (protons) generated by the oxidation of water help create a proton gradient that is used by ATP synthase to generate ATP. Protons are the terminal acceptors of these photosynthetically generated electrons in the algal chloroplast. The process results in the simultaneous produc­tion of oxygen and hydrogen gases (Spruit, 1958; Greenbaum et al., 1983).

Direct photolysis capitalizes on the photosynthetic capability of microalgae and cyanobacteria to split water directly into oxygen and hydrogen. Cyanobacteria, also known as blue-green algae, belong to a phylum of bacteria that obtain their energy through photo­synthesis. Microalgae have evolved the ability to harness solar energy by extracting protons

and electrons from water via water-splitting reactions. The biohydrogen production takes place via direct absorption of light and transfer of electrons to two groups of enzymes: hydrogenases and nitrogenases (Manis and Banerjee, 2008). Under anaerobic conditions or when too much energy is captured in the process, some microorganisms vent the excess electrons by using a hydrogenase enzyme that converts the hydrogen ions to hydrogen gas (Sorensen, 2005; Turner et al., 2008). It has been reported that the protons and electrons extracted via the water-splitting process are recombined by a chloroplast hydrogenase to form molecular hydrogen gas with a purity of up to 98% (Hankamer et al., 2007).

In addition to producing hydrogen, the microorganisms also produce oxygen, which in turn suppresses hydrogen production (Kovacs et al., 2006; Kapdan and Kargi, 2006). Research work has been carried out to engineer algae and bacteria so that the majority of the solar energy is diverted to hydrogen production, with bare energy diverted to carbohydrate production to solely maintain cells. Researchers are attempting to either identify or engineer less oxygen-sensitive microorganisms, isolate the hydrogen and oxygen cycles, or change the ratio of photosynthesis to respiration to prevent oxygen buildup (U. S. DOE, 2007). Addition of sulfate has been found to suppress oxygen production. However, the hydrogen production mechanisms are also inhibited (Sorensen, 2005; Turner et al., 2008).

The merit of direct photolysis is that the principal feed is water and the driver energy is derived from sunlight, both of which are readily available. Although this technology has significant promise, it is also facing tremendous challenges. A major challenge is the incom­patibility in the simultaneous molecular hydrogen and oxygen production. Photosynthetic hydrogen can only be produced transiently, since oxygen is a strong suppressor of hydroge­nate reactions and a powerful inhibitor of the [Fe]-hydrogenase. In addition, the photolysis process requires a significant algae cultivation area to collect sufficient light. Another challenge is achieving continuous hydrogen production under aerobic conditions (U. S. DOE, 2007).