Full details on continuous biohydrogen production from sugarcane molasses are unraveled: Performance optimization, self-regulation, metabolic correlations and quanti-qualitative biomass characterization

Abstract Using pure sugars for fermentative biohydrogen (bioH2) production is known as an economically impeditive approach due to the high costs of raw materials, which leads to the frequent exploitation of residual streams. However, the potentials of using biodigestion as a core processing step in sugarcane biorefineries opens up a wide range of biotechnological possibilities, in which sugar-rich materials may be fermented without adding acquisition costs. This study aimed to finalize the full optimization of the continuous long-term (630 d) thermophilic (55 °C) bioH2 production from molasses, defining adequate hydraulic retention time (HRT) levels. Details of temporal and spatial metabolite distribution profiles coupled to the characterization of microbial communities provided the most complete picture of molasses acidogenesis to date. BioH2 evolution (8.5 NL-H2 L−1 d−1) in full optimized conditions, i.e., HRT ≈ 10.0 h, organic loading rate (OLR) ≈ 86.0 kg-CODt m−3 d−1 and pH ≈ 5.38, exceeded the individual application of optimal OLR (4.5 NL-H2 L−1 d−1) and pH (2.4 NL-H2 L−1 d−1) by almost 200% and 350%, respectively. Biomass washout naturally controlled substrate availability (6.21 ± 2.1 g-COD g−1VSS d−1 for over 300 d), preventing performance losses in the long-term. BioH2 derived from acetic- and butyric-type fermentations was observed in the basal portion of the reactor, whilst butyrate and bioH2 from reverse β-oxidation using lactate prevailed in the bed region. The Thermoanaerobacterium genus was the main group involved in such pathways, and the contribution of the Caproiciproducens genus in caproate production was also revealed. These findings provide consistent directions to scale-up acidogenic systems towards an effective exploitation of full-scale bioH2 production.