Investigation of Integrated Hydrogen Production Through the Coupling of Dark Fermentation, Photofermentation, and Microbial Electrolysis, project financed by National Science Centre, project leader: prof. Stanisław Ledakowicz

Biohydrogen (bioH₂) has emerged as a key element of the transition toward sustainable energy systems due to its high energy density and clean combustion, producing only water as a by-product. It can be generated through dark fermentation (DF), photofermentation (PF), or microbial electrolysis cells (MECs). Among these routes, DF is particularly attractive because of its flexibility toward organic substrates and its capacity to valorise food and biowaste. DF of kitchen waste has proven to be an efficient and robust biological pathway in our previous studies, where stable bioH₂ production was achieved under controlled conditions. However, DF is limited by the accumulation of volatile fatty acids (VFAs) and other metabolites that inhibit microbial activity and constrain hydrogen yield. Consequently, additional downstream processes are required to convert these intermediates and enhance overall bioH₂ recovery. This project aims to develop an integrated valorisation strategy for dark fermentation digestate, focusing on the liquid fraction while considering the entire digestate stream within a circular biorefinery framework. The liquid phase will be processed through photofermentation and microbial electrolysis clls (MECs) to recover additional hydrogen from VFAs and other residual organics. PF uses purple non-sulfur photosynthetic bacteria under illuminated, photoheterotrophic conditions to convert VFAs into hydrogen, and the choice of photobioreactor configuration, adjustment of photo-bioprocess parameters kinetics of bacterial growth coupled with H₂ production rate and potential use of photocatalysts as well as scaling-up represent important research challenges.

MECs have emerged as a promising technology for sustainably producing bioH₂ from organic wastes. MECs can generate H₂ at higher yields using VFAs. MECs, in turn, provide a bioelectrochemical route for bioH₂ production supported by electroactive consortia and enzymatic catalysts such as hydrogenases. However, uncertainties remain regarding reactor design (single- vs two-chamber systems), optimal electrode and membrane materials, and the coupling of DF with MECs, as most existing studies rely on single carbon sources rather than real digestates. The DF-MEC system effectively overcomes the limitations of conventional MECs, such as high voltage requirements and low hydrogen yields, by using the VFAs and other byproducts generated during DF as substrates for MEC. Key parameters to explore include evaluating different substrates, optimizing hydraulic retention time (HRT), and understanding the combined effects of pH and substrate loading on H₂ generation.

Because DF produces effluents with high turbidity and fine solids that hinder light penetration and promote membrane fouling, the project will begin with the investigation of pretreatment and clarification procedures to ensure suitable feedstock quality for PF and MEC. The influence of substrate type - complex kitchen waste versus simpler lignocellulosic material such as corn stalks - on digestate composition, inhibitory effects, and downstream process performance will be investigated. The solid fraction of the digestate will be further directed to methane fermentation to maximize carbon utilisation, while the feasibility of recycling liquid effluents after PF, MEC, or their combination back into the DF step as dilution water will be evaluated. These recycling routes will not constitute the main research focus but will provide insight into the potential circularity of the integrated system. A key innovative aspect of this work is the investigation of DF+PF, DF+MEC, and the l unexplored three-stage DF+PF+MEC hybrid configuration, in which each process utilises fermentation intermediates not metabolized in the previous step. Such integration could significantly increase the total bioH₂ yield while enabling simultaneous treatment of biowaste streams. Furthermore, the project will assess the energy efficiency and mass–energy balance of the integrated configurations, an area currently absent in the literature despite its importance for evaluating the suitability and competitiveness of bioH₂ systems. By linking DF with PF and/or MEC into a coherent bioprocess chain, the project is expected to enhance bioH₂ production, minimize waste discharge, and promote nutrient and water circularity. Ultimately, the results will contribute to the development of energy-positive, zero-waste bioH₂ biorefineries.