Dark fermentative hydrogen production by the unicellular halotolerant cyanobacterium Aphanothece halophytica grown in seawater

Biohydrogen is an environmentally friendly alternative energy carrier that can be produced by a number of different microorganisms. The unicellular halotolerant cyanobacterium Aphanothece halophytica is one of the high potential H-2 producers. Under dark fermentation, it is capable of producing H-2 by the bidirectional hydrogenase activity via the catabolism of glycogen stored during photosynthesis. This work aimed to cultivate A. halophytica in natural seawater containing high salinity and minerals, with an addition of some essential nutrients, and to investigate effects of various nutritional and physical factors on its dark fermentative H-2 production. A. halophytica was able to grow in natural seawater added with NaNO3. Cells grown in seawater supplemented with as little as 1.76 mM NaNO3 showed similar growth to those cultivated in normal BG11 supplemented with Turk Island salt solution. H-2 production was the highest when incubating the cells in seawater without any supplementation of NaNO3. Under this condition, the highest rate of dark fermentative H-2 production of 82.79 +/- 3.47 nmol H-2 mg(-1) dry weight h(-1) was found in cells incubated at 35 A degrees C, pH 6 with the supplementation of 378 mmolC L-1 glucose, 0.25 M NaCl, and 0.4 mu M Fe3+. Long-term H-2 accumulation of 1,864 A +/- 81 nmol H-2 mg(-1) dry weight was observed after 8 days of dark incubation under anoxic condition, and the high yield of H-2 was sustained at least up to 14 days, suggesting the possibility of utilizing natural seawater to grow A. halophytica for long-term production of H-2.