Flocculation-enhanced photobiological hydrogen production by microalgae: Flocculant composition, hydrogenase activity and response mechanism

The practical application of microalgal photobiological hydrogen was limited by its low yield and short production time. In this study, the promotion of inorganic flocculants (polymeric aluminum chloride, PAC) and organic flocculants (cationic polyacrylamide, CPAM) to the aggregate formation and photobiological hydrogen production of prokaryotic microalgae (Synechocystis sp.) and eukaryotic microalgae (Chlamydomonas reinhardtii) were investigated. The effectiveness of PAC-CPAM composite flocculants in promoting aggregation was compared to single PAC or CPAM, with flocculation efficiencies exceeding 92.2 % for both microalgal strains. After aggregate formation, hydrogen production in Synechocystis and C. reinhardtii increased by 65.9 % and 80.2 %, respectively, compared with the control (without flocculation). Their hydrogenase activity rose to 1.14 and 28.2 μmol H2/(mg chlorophyll·h), respectively. Aggregation sustained the photosynthetic activity of both Synechocystis and C. reinhardtii, with the photosynthetic system electron transfer rate (ETR) 1.54–1.72 times higher than that of the control. The formation of aggregates enhanced microalgal respiration, accelerating the reduction of oxygen content, and increasing hydrogenase activity. Notably, the increased respiration in Synechocystis and C. reinhardtii aggregates supplied more electrons, enhancing hydrogen production by consuming intracellular organic matter. Aggregate formation upregulated the expression of Synechocystis and C. reinhardtii hydrogenase genes by average 2.1-fold and 2.5-fold, respectively. Simultaneously, the expression of genes related to photosynthetic system Ⅱ, ferredoxin, pyruvate ferredoxin-oxidoreductase, and oxidative phosphorylation were upregulated, with a significant effect observed in C. reinhardtii aggregates. The results of this study advanced the utilization and industrialization of microalgae-based green hydrogen production, contributing to carbon neutrality.