Sulfate- and pH-driven metabolic flexibility in sugarcane vinasse dark fermentation stimulates biohydrogen evolution, sulfidogenesis or homoacetogenesis

Abstract

Dark fermentation of sugarcane vinasse can be used as a “cleaning” step to remove sulfate prior to methanogenesis because sulfidogenic conditions can be successfully established in parallel with biohydrogen production. Using a 22 central composite rotational design (CCRD) and response surface methodology (RSM), this study assessed the impacts of bicarbonate and sulfate availability on the establishment of sulfidogenesis in the thermophilic (55 °C) fermentation of vinasse in batch reactors, equally assessing the impacts on biohydrogen evolution. CCRD-RSM results indicated the favoring of biohydrogen production at the lowest sulfate and bicarbonate concentrations, whilst the opposite was observed for sulfidogenesis. Glycerol, lactate, and hydrogen were the preferential electron donors utilized by sulfate-reducing bacteria (SRB), whilst ethanol was markedly consumed only at high sulfate concentrations. SRB were inhibited by sodium when dosing excess NaHCO3 and Na2SO4. Complementary tests revealed maximum biohydrogen production (2.40 mmol) out of the CCRD, at pH exceeding 7.5 with no interference of sulfidogenesis. Non-efficient biohydrogen production was observed at low pH (<5.0; ∼1.90 mmol) because the uptake of lactate was inhibited. Meanwhile, homoacetogenesis was established under intermediate pH range (5.5–6.5), as revealed by the accumulation of acetate (up to 2.5 g L−1). 16S rRNA gene amplicon sequencing further revealed the genera Thermoanaerobacterium/Pseudoclostridium, Desulfotomaculum/Desulfohalotomaculum and Sporomusaceae/Moorella as the main biohydrogen-producing, sulfate-removing and biohydrogen-consuming (homoacetogens) microbial groups, respectively. Hence, using a single inoculum source, vinasse may provide a butyrate-rich (along with biohydrogen-rich biogas) or a sulfate-free and acetate-rich fermented effluent, depending mainly on proper pH control.