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Anaerobic digestion (AD) is a well-established technology used for producing biogas or biomethane alongside the slurry used as biofertilizer. However, using a variety of wastes and residuals as substrate and mixed cultures in the bioreactor makes AD as one of the most complicated biochemical processes employing hydrolytic, acidogenic, hydrogen-producing, acetate-forming bacteria as well as acetoclastic and hydrogenoclastic methanogens. Hydrogen and volatile fatty acids (VFAs) including acetic, propionic, isobutyric, butyric, isovaleric, valeric and caproic acid and other carboxylic acids such as succinic and lactic acids are formed as intermediate products. As these acids are important precursors for various industries as mixed or purified chemicals, the AD process can be bioengineered to produce VFAs alongside hydrogen and therefore biogas plants can become biorefineries. The current critical review paper provides the theory and means to produce and accumulate VFAs and hydrogen, inhibit their conversion to methane and to extract them as the final products. The effects of pretreatment, pH, temperature, hydraulic retention time (HRT), organic loading rate (OLR), chemical methane inhibitions, and heat shocking of the inoculum on VFAs accumulation, hydrogen production, VFAs composition, and the microbial community were discussed. Furthermore, this paper highlights the possible techniques for recovery of VFAs from the fermentation media in order to minimize product inhibition as well as to supply the carboxylates for downstream procedures.

Citrus waste is a promising potential feedstock for anaerobic digestion, yet the presence of inhibitors such as d-limonene is known to limit the process. Effluent recirculation has been proven to increase methane yield in a semi-continuous process for recalcitrant material, but it has never been applied to toxic materials. This study was aimed to investigate the effect of recirculation on biogas production from citrus waste as toxic feedstock in two-stage anaerobic digestion. The first digestion was carried out in a stirred tank reactor (STR). The effluent from the first-stage was filtered using a rotary drum filter to separate the solid and the liquid phase. The solid phase, rich in hydrophobic D-limonene, was discarded, and the liquid phase containing less D-limonene was fed into the second digester in an up-flow anaerobic sludge bed (UASB) reactor. A high organic loading rate (OLR 5 g VS/(L·day)) of citrus waste was fed into the first-stage reactor every day. The effluent of the first-stage was then fed into the second-stage reactor. This experiment was run for 120 days. A reactor configuration without recirculation was used as control. The result shows that the reactor with effluent recirculation produced a higher methane yield (160–203 NmL/g·VS) compared to that without recirculation (66–113 NmL/g·VS). More stable performance was also observed in the reactor with recirculation as shown by the pH of 5–6, while without recirculation the pH dropped to the range of 3.7–4.7. The VS reduction for the reactor with recirculation was 33–35% higher than that of the control without recirculation. Recirculation might affect the hydrolysis-acidogenesis process by regulating pH in the first-stage and removing most of the D-limonene content from the substrate through filtration.

The presence of an antimicrobial compound called D-Limonene in citrus waste inhibits methane production from such waste in anaerobic digestion. In this work, a two-stage anaerobic digestion method is developed using reverse membrane bioreactors (rMBRs) containing cells encased in hydrophilic membranes. The purpose of encasement is to retain a high cell concentration inside the bioreactor. The effectiveness of rMBRs in reducing cell washout is evaluated. Three different system configurations, comprising rMBRs, freely suspended cells (FCs), and a combination of both (abbreviated to rMBR–FCs), are incubated at three different organic loading rates (OLRs) each, namely 0.6, 1.2, and 3.6 g COD/(L cycle). Incubation lasts for eight feeding cycles at 55 °C. Methane yield and biogas composition results show that rMBRs perform better than rMBR–FCs and FCs at all three OLRs. Volatile fatty acid profiles and H2 production show that the reactors are working properly and no upset occurs. Additionally, a short digestion time of 4 days can be achieved using the rMBR configuration in this study.

Anaerobic digestion is a promising method to treat citrus waste. However, the presence of limonene in citrus waste inhibits anaerobic digestion process. Limonene is an antimicrobial compound and could inhibit methane forming bacteria that takes a longer time to recover than the injured acid forming bacteria. Hence, volatile fatty acids will be accumulated and methane production will be decreased. One way to solve this problem is by conducting anaerobic digestion process into two stages. The first step is aimed for hydrolysis, acidogenesis, and acetogenesis reactions and the second stage is aimed for methanogenesis reaction. The separation of the system would further allow each stage in their optimum conditions making the process more stable. In this research, anaerobic digestion was carried out in batch operations using 120 ml-glass bottle bioreactors in 2 stages. The first stage was performed in free-cells bioreactor, whereas the second stage was performed in both bioreactor of free cells and membrane bioreactor. In the first stage, the reactor was set into 'anaerobic' and 'semi-aerobic' conditions to examine the effect of oxygen on facultative anaerobic bacteria in acid production. In the second stage, the protection of membrane towards the cells against limonene was tested. For the first stage, the basal medium was prepared with 1.5 g VS of inoculum and 4.5 g VS of citrus waste. The digestion process was carried out at 55°C for four days. For the second stage, the membrane bioreactor was prepared with 3 g of cells that were encased and sealed in a 3×6 cm2polyvinylidene fluoride membrane. The medium contained 40 ml basal medium and 10 ml liquid from the first stage. The bioreactors were incubated at 55°C for 2 days under anaerobic condition. The results from the first stage showed that the maximum total sugar under 'anaerobic' and 'semi-aerobic' conditions was 294.3 g/l and 244.7 g/l, respectively. The corresponding values for total volatile fatty acids were 3.8 g