Improvement of H2 photoproduction in Chlorella pyrenoidosa in artificial and natural seawater by addition of acetic acid and control of nutrients

•Acetic acid greatly promoted H2 production by marine Chlorella in ASW medium.•S and N deprivation further enhanced H2 production by marine Chlorella.•C. pyrenoidosa 707S evolved H2 in natural seawater medium under N deprivation.

Two marine Chlorella strains, Chlorella pyrenoidosa 689S and C. pyrenoidosa 707S, were previously found to generate H2 gas under sulfur (S) deprivation. In this work, their ability to generate H2 under sulfur-replete, artificial and natural seawater conditions was further explored. C. pyrenoidosa 689S and 707S evolved 20.77 and 25.51 ml/l H2, respectively, in artificial seawater (ASW) without any nutrient deprivation, simply by adding acetic acid. Sulfur or nitrogen (N) deprivation promoted H2 production by C. pyrenoidosa 689S and 707S incubated in ASW, while phosphorous (P) deprivation did not improve H2 production effectively. Changes of culture medium from ASW-S to TAP′-S induced an increase of ~ 14% and 200% in H2 yield in C. pyrenoidosa 689S and 707S respectively. However, the Chlorella cells did not grow well in TAP′ medium, which contains NH4Cl as nitrogen source, at approximately the same molar concentration as the nitrogen in ASW medium, as indicated by the low chlorophyll content in TAP′-S cultures. When natural seawater was used as the water source for culture medium instead of the ASW prepared using distilled water, the H2 production by C. pyrenoidosa 689S and 707S was significantly altered. Both Chlorella strains failed to transit to the anaerobic phase and evolved no H2 in L1 medium containing acetate prepared using natural seawater, implying that some unknown factor(s) in natural seawater might hinder the establishment of anaerobiosis and activation of H2ase. Even N and P deprivation did not induce H2 production in C. pyrenoidosa 689S. Although C. pyrenoidosa 707S was not able to evolve H2 under P-deprivation conditions in natural seawater medium, it was found, surprisingly, to generate comparable amounts of H2 in N-free medium prepared using natural seawater as in N-free ASW medium, showing that the strain might be useful for biological H2 photoproduction.