1992). Thus, to check whether C. reinhardtii cells are already in the stage of S deprivation, 1 ml of the cells is removed from the culture vessel and mixed with 10 μl of a 30 mM XSO4 stock solution in 0.1 M Tris/HCl pH 7.5. After 30–60 min, the cells are spun down at a high speed, and the supernatant, which should be visibly bluish if an arylsulfatase is active, can be analyzed photospectrometrically

at λ = 650 nm. In BIBF 1120 solubility dmso contrast to the simplicity of inducing S starvation in C. reinhardtii, the induction of a sustained and reproducible H2 production in these cultures is much more difficult. To understand this difficulty, the sequence of events leading to the onset of H2 production in C. reinhardtii is briefly summarized here. When the cells have been transferred to S-free medium and placed in the light, they still have a high photosynthetic activity, resulting in O2 selleck evolution and Rabusertib mw CO2 fixation. The latter results not only in some cell growth and doubling in the beginning (Melis et al. 2000), but also in the accumulation of starch, which is a common response of nutrient starved C. reinhardtii cells (Grossman 2000). Starch levels had tripled already

in the first 5 h of S depletion (Makarova et al. 2007), and after being S depleted for 24 h, the algae contain almost tenfold amounts of starch as compared with S-replete cells (Zhang et al. 2002). After several hours, PSII activity and photosynthetic O2 evolution, respectively, as well as CO2 fixation will decrease (Melis et al. 2000; Hemschemeier et al. 2008). At Cetuximab manufacturer a certain point, the

ongoing respiratory O2 uptake activity will overcome photosynthetic O2 evolution rates so that the O2 dissolved in the culture will be consumed by and by (Fig. 3). As soon as anaerobic conditions are established, the hydrogenase gene is expressed (Zhang et al. 2002) and the hydrogenase enzyme becomes active (Winkler et al. 2002b) (Fig. 3). The hydrogenase then takes over the electrons from ferredoxin, which in turn is reduced by PSI activity. The electrons arriving at PSI originate both from residual PSII activity and non-photochemical PQ-reduction (Fouchard et al. 2005; Hemschemeier et al. 2008) (Fig. 1b). The latter, again, depends on the amount of starch that was accumulated in the photosynthetic phase. Fig. 3 a Development of the concentrations of H2 (●), O2 ( ), and CO2 (○) as measured by MS in the headspace of an S-depleted C. reinhardtii culture incubated in squared glass bottles sealed with Suba seals upon one-site illumination as illustrated by the photograph in (b) (Hemschemeier 2005) Having these metabolic adaptations of S-depleted algae under consideration, it is obvious that every culture parameter influencing the photosynthetic light and dark reactions as well as respiratory activity will also have an impact on establishment and dimension of the photosynthetic H2 metabolism.