Non-conventional pretreatments mitigate the inhibitory effect of 5-hydroxymethylfurfural in dark fermentation process

Dark fermentation carried out by mixed microbial cultures is considered a promising process for the production H2 biofuel. However, this process is still not cost effective and unreliable. In mixed culture based systems, the H2 production is enhanced by pretreating the initial seed to suppress the H2 consumers activity. However, this approach has been reported to influence the structure and diversity of H2 producing bacteria (HPB), which may have negative effects on the resilience and resistance of the system when changes of environmental conditions, or presence of inhibitors, occurs. 2. Objectives This study evaluates the effect of pretreatments on the performance and structure of the enriched microbial community and its potential response to an inhibitor (5-hydroxymethylfurfural, HMF). All systems produced hydrogen with a yield ranging between 0.6-0.9 mol H2/mol glucose. HMF decreased the H2 production of 19.1±2.9 % in most of the systems. However, in the pre-aerated systems the hydrogen potential was 22% higher than the other systems without significant difference between C and HMF reactors. Extended lag phases were observed in the pre-acidified tests, though the discrepancy of the lag time in the C and HMF tests was smaller than the other bottles. The high performance of the pre-aerated and acidified HMF was due to increased metabolic activity of the main taxa observed in the systems (Paenibacillus and Clostridium spp.) rather than significant changes in the community composition. In conclusion, pre-aeration and acidification of the initial inoculum can mitigate the negative effect of the HMF. The stressful conditions applied to the original inoculum may have activated cell/community response mechanisms able to counteract the toxicity of the HMF.