Abstract
The unicellular halotolerant cyanobacterium Aphanothece halophytica is a potential dark fermentative producer of molecular hydrogen (H(2)) that produces very little H(2) under illumination. One factor limiting the H(2) photoproduction of this cyanobacterium is an inhibition of bidirectional hydrogenase activity by oxygen (O(2)) obtained from splitting water molecules via photosystem II activity. The present study aimed to investigate the effects of the photosystem II inhibitors carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) on H(2) production of A. halophytica under light and dark conditions and on photosynthetic and respiratory activities. The results showed that A. halophytica treated with CCCP and DCMU produced H(2) at three to five times the rate of untreated cells, when exposed to light. The highest H(2) photoproduction rates, 2.26 ± 0.24 and 3.63 ± 0.26 μmol H(2 )g(-1) dry weight h(-1), were found in cells treated with 0.5 μM CCCP and 50 μM DCMU, respectively. Without inhibitor treatment, A. halophytica incubated in the dark showed a significant increase in H(2) production compared with cells that were incubated in the light. Only CCCP treatment increased H(2) production of A. halophytica during dark incubation, because CCCP functions as an uncoupling agent of oxidative phosphorylation. The highest dark fermentative H(2) production rate of 39.50 ± 2.13 μmol H(2 )g(-1) dry weight h(-1) was found in cells treated with 0.5 μM CCCP after 2 h of dark incubation. Under illumination, CCCP and DCMU inhibited chlorophyll fluorescence, resulting in a low level of O(2), which promoted bidirectional hydrogenase activity in A. halophytica cells. In addition, only CCCP enhanced the respiration rate, further reducing the O(2) level. In contrast, DCMU reduced the respiration rate in A. halophytica.