Hydrogen production by photoautotrophic sulfur-deprived Chlamydomonas reinhardtii pre-grown and incubated under high light.

We have previously demonstrated that Chlamydomonas reinhardtii can produce hydrogen under strictly photoautotrophic conditions during sulfur deprivation [Tsygankov et al. (2006); Int J Hydrogen Energy 3:1574-1584]. The maximum hydrogen photoproduction was achieved by photoautotrophic cultures pre-grown under a low light regime (25 microE m(-2) s(-1)). We failed to establish sustained hydrogen production from cultures pre-grown under high light (100 microE m(-2) s(-1)). A new approach for sustained hydrogen production by these cultures is presented here. Assuming that stable and reproducible transition to anerobiosis as well as high starch accumulation are important for hydrogen production, the influence of light intensity and dissolved oxygen concentration during the oxygen evolving stage of sulfur deprivation were investigated in cultures pre-grown under high light. Results showed that light higher than 175 microE m(-2) s(-1) during sulfur deprivation induced reproducible transition to anerobiosis, although the total amount of starch accumulation and hydrogen production were insignificant. The potential PSII activity measured in the presence of an artificial electron acceptor (DCBQ) and an inhibitor of electron transport (DBMIB) did not change in cultures pre-grown under 20 microE m(-2) s(-1) and incubated under 150 microE m(-2) s(-1) during sulfur deprivation. In contrast, the potential PSII activity decreased in cultures pre-grown under 100 microE m(-2) s(-1) and incubated under 420 microE m(-2) s(-1). This indicates that cultures grown under higher light experience irreversible inhibition of PSII in addition to reversible down regulation. High dissolved O(2) content during the oxygen evolving stage of sulfur deprivation has a negative regulatory role on PSII activity. To increase hydrogen production by C. reinhardtii pre-grown under 100 microE m(-2) s(-1), cultures were incubated under elevated PFD and decreased oxygen pressure during the oxygen evolving stage. These cultures reproducibly reached anaerobic stage, accumulated significant quantities of starch and produced significant quantities of H(2). It was found that elevation of pH from 7.4 to 7.7 during the oxygen producing stage of sulfur deprivation led to a significant increase of accumulated starch. Thus, control of pH during sulfur deprivation is a possible way to further optimize hydrogen production by photoautotrophic cultures.

Hydrogen photoproduction by nutrient-deprived Chlamydomonas reinhardtii cells immobilized within thin alginate films under aerobic and anaerobic conditions.

A new technique for immobilizing H2-photoproducing green algae within a thin (<400 microm) alginate film has been developed. Alginate films with entrapped sulfur/phosphorus-deprived Chlamydomonas reinhardtii, strain cc124, cells demonstrate (a) higher cell density (up to 2,000 microg Chl mL(-1) of matrix), (b) kinetics of H2 photoproduction similar to sulfur-deprived suspension cultures, (c) higher specific rates (up to 12.5 micromol mg(-1) Chl h(-1)) of H2 evolution, (d) light conversion efficiencies to H2 of over 1% and (e) unexpectedly high resistance of the H2-photoproducing system to inactivation by atmospheric O2. The algal cells, entrapped in alginate and then placed in vials containing 21% O2 in the headspace, evolved up to 67% of the H2 gas produced under anaerobic conditions. The results indicate that the lower susceptibility of the immobilized algal H2-producing system to inactivation by O2 depends on two factors: (a) the presence of acetate in the medium, which supports higher rates of respiration and (b) the capability of the alginate polymer itself to effectively separate the entrapped cells from O2 in the liquid and headspace and restrict O2 diffusion into the matrix. The strategy presented for immobilizing algal cells within thin polymeric matrices shows the potential for scale-up and possible future applications.

Prolongation of H2 photoproduction by immobilized, sulfur-limited Chlamydomonas reinhardtii cultures.

Two approaches to prolong the duration of hydrogen production by immobilized, sulfur-limited Chlamydomonas reinhardtii cells are examined. The results demonstrate that continuous H2 photoproduction can occur for at least 90 days under constant flow of TAP medium containing micromolar sulfate concentrations. Furthermore, it is also possible to prolong the duration of H2 production by cycling immobilized cells between minus and plus sulfate conditions.