Hydrogen and Methane Generation from Biowaste: Enhancement and Upgrading via Bioelectrochemical Systems

Abstract

Bioelectrochemical systems (BESs) are emerging technologies that are based on catalyzing (bio-)anode and (bio-)cathode reactions from waste biomass by exoelectrogenic microorganisms. Microbial electrolysis cell (MEC), which is one of the BESs’ technologies, is typically used to degrade organic wastes or wastewater for bioenergy recovery and biosynthesis. As one of the promising biotechnologies for resource recovery, value-added products have been obtained by MEC- or ME-integrated systems, such as hydrogen, methane, ethanol, etc. The fundamental reactions of (bio-)electron transport through anodic oxidation are well understood and allow us to increase reactor performance and efficiency. More attentions have been recently paid to cathode reactions on proton/electron transport and recovery, with or without microbial activities. Biogas upgrading systems have also been promoted in integrated systems, by combining bioelectrochemistry with various anaerobic processes. This chapter will focus on energy gas generation from waste organics involved in bioelectrochemical pathways and give an overview of bottlenecks and challenges related to this technology.

5.1 Principle for Hydrogen and Methane Generation via Bioelectrochemical Systems

With the continuous stimulation of fossil fuel consumption and energy demand growth, the need to combine energy security with the development of a more sustainable energy sector is representing a severe global challenge. Some renewable resources are considered promising to harvest clean energy. Hydrogen is in principle an environmentally acceptable and clean energy vector that, however, is typically produced from non-renewable fossil fuels such as natural gas or water. In theory, bioelectrochemical systems (BESs) can be used to recover hydrogen from any biodegradable organics on the (bio-) cathode, by harnessing the biocatalytic electrolysis (bio-) anode, i.e., microbial electrolysis cell (MEC). Biocatalytic oxidation of organic matter to hydrogen can occur at both the anode and the cathode. In MEC, for instance, biodegradable organics can produce electrons through the biological oxidation at the anode. Electrons flow then through an external circuit (going from the anode to the cathode) and subsequently combine with protons to form hydrogen under a small voltage (0.2–1.2 V), necessary to overcome thermodynamic barriers (Cheng and Logan 2007). Additionally, H₂, being one of the most effective electron shuttles, can be exploited by microorganisms to produce small molecular compounds such as methane (Fig. 5.1). Compared with other H₂-/CH₄-producing technologies, MEC, as bioelectrochemical power-to-gas, which can convert renewable surplus electricity into hydrogen and methane, exhibits higher H₂-/CH₄-producing efficiency and wider diversity of substrate utilization, making it more advantageous, especially for the valorization of low concentration and/or complex organic matter (Logan et al. 2008).

Fig. 5.1

Schematic of H₂ and CH₄ formation in the microbial electrolysis cell, MEC

5.1.1 Reactions Based on Extracellular Electron Transport

Based on whether there are microorganisms attached on the electrodes or not, MEC can be divided into four categories: (1) full-biological double-chambered (DC) bioanode/biocathode MEC; (2) full-biological single-chambered (SC) bioanode/biocathode MEC; (3) half-biological double-chambered bioanode/cathode MEC; and (4) half-biological double-chambered anode/biocathode MEC.

In general, biodegradable organic matter is oxidized by anodic exoelectrogenic microorganisms and releases protons and carbon dioxide into the anolyte, while electrons transfer to the anode in a typical dual-chamber MEC (Hamelers et al. 2010).

[Anodic reaction]

$$ {\mathrm{CH}}_3\mathrm{COOH}+{2\mathrm{H}}_2\mathrm{O}\to {2\mathrm{CO}}_2+{8\mathrm{H}}^{+}+{8\mathrm{e}}^{-},{E}^0=0.187\ \mathrm{and}\ E=-0.289\ \mathrm{V}\ \mathrm{vs}.\mathrm{SHE} $$

(5.1)

Electrons flow typically through an external circuit, driven by an external voltage. Protons and carbon dioxide diffuse across the separator (such as ion exchange membranes, size-selective membranes, stacks of membranes, or the cloth) to the cathode, combining with electrons to be reduced to hydrogen or methane, depending on the cathodic potential, which drives the H₂/CH₄ production, affecting Gibbs’ free energy (Jafary et al. 2015).

[Cathodic reaction]

Hydrogen formation:

$$ {8\mathrm{H}}^{+}+{8\mathrm{e}}^{-}\to {4\mathrm{H}}_2,{E}^0=0\ \mathrm{and}\ E=-0.412\ \mathrm{V}\ \mathrm{vs}.\mathrm{SHE} $$

(5.2)

Methane formation:

$$ {\mathrm{CO}}_2+{4\mathrm{H}}_2\to {\mathrm{CH}}_4+{2\mathrm{H}}_2\mathrm{O},{E}^0=0.227\ \mathrm{V}\ \mathrm{and}\ E=-0.248\ \mathrm{V}\ \mathrm{vs}.\mathrm{SHE} $$

(5.3)

Microbial electrolysis for hydrogen production can be considered as an advantageous combination of conventional hydrogen production pathways. First, compared to dark fermentation, “hydrogen-producing microorganisms” of MEC cannot directly produce hydrogen but are capable of extracellular electron transfer, which play a pivotal role. Besides, MEC can also handle more types of organic substrates without the problem of fermentation end products, which provides a possible way to thoroughly and fully degrade organic matter. In addition, the efficiency of hydrogen production via dark fermentation (such as treated carbohydrate-rich wastewater) is limited (Angenent et al. 2004), due to thermodynamical limitations involving endothermic reactions (Hawkes et al. 2002; Kim et al. 2004; Oh et al. 2003). Second, compared with photosynthetic biological hydrogen production (photosynthesis, photo-fermentation), it is not restricted by light. Notably, photo-fermentation requires enormous reactor surface areas to improve electron transfer and overcome the diffuse nature of solar radiation and thermodynamical barriers, which are clearly not economically viable (Hallenbeck and Benemann 2002). Third, the cathode reaction has the same reaction as the electrolysis of hydrogen in the electrochemical method, except that the electron source of the anode is different. As a consequence, the electrons provided by the relevant microorganisms can save input energy, as demonstrated by the minimum applied potential. The theoretical minimum potential of hydrogen production reaction in MEC is 0.10 V (shown in the following, Eqs. (5.4) and (5.5)), whereas the theoretical minimum potential of hydrogen production in industrial electrolysis is 1.2 V.

For the hydrogen production in MEC, the reaction between proton and electron occurs only in the cathode; the semi-cell reaction of its electrochemical system is briefly described as:

$$ \mathrm{Anodic}\ \mathrm{reaction}:{\mathrm{NAD}}^{+}+{\mathrm{H}}^{+}+{2\mathrm{e}}^{-}\to \mathrm{NADH},{E}^0=-0.320\ \mathrm{V}\ \mathrm{vs}.\mathrm{SHE} $$

(5.4)

$$ \mathrm{Cathodic}\ \mathrm{reaction}:{2\mathrm{H}}^{+}+{2\mathrm{e}}^{-}\to {\mathrm{H}}_2,{E}^0=-0.420\ \mathrm{V}\ \mathrm{vs}.\mathrm{SHE} $$

(5.5)

To enable a nonspontaneous reaction, electrons are required to flow from the anode potential (−0.320 V, Eq. (5.4)) to the cathode potential (−0.420 V, Eq. (5.5)), namely, from the high point to the low point. Thereby, MEC needs extra energy to execute the reverse flow of electrons. Theoretically, this process needs to provide a potential of at least 0.1 V to overcome the energy barrier, whereas in practice, an additional voltage supply of minimum 0.13 V is needed to perform the cathode hydrogen production. The reason for the extra voltage required is that electrochemically active microorganisms or electroactive microorganisms (EAMs) consume some of available energy to sustain their own growth, resulting in microbes to release electrons with a higher potential than the equilibrium potential. On the other hand, the voltage applied to the reactor will also be lost as the consequences of ohmic resistance of electrochemical bias and occurrence of overpotential (Rozendal et al. 2006). Nevertheless, hydrogen generated through water electrolysis actually requires at least a voltage of 1.6 V or more (Crow 1994; Rasten et al. 2003).

5.1.2 Functional Communities Involved in Bioelectrochemical Systems

The term EAMs refers to those microorganisms that can directly or indirectly donate electrons to an electrode (called exoelectrogens), or that accept electrons from the electrode (known as electrotrophs) (Table 5.1). Thus far, the exoelectrogens isolated from natural environment belong primarily to the phylum Proteobacteria and Firmicutes, mostly facultative anaerobic microorganisms, which are capable of gaining energy to sustain growth via anaerobic respiration and fermentation metabolism. Most exoelectrogens are Fe(III)-reducing bacteria (FRB), viz., the oxidized iron is the final electron acceptor of the respiratory chain (Lovley 2006). There are different strategies for transferring electrons to the anode, such as the mediated interspecies electron transfer (MIET) (Cai et al. 2020) and the direct interspecies electron transfer (DIET) (Logan et al. 2019; Lovley 2017). The latter requires direct contact of the outer membrane cytochromes and electron transport proteins associated with outer cell surfaces on electrically conductive materials. MIET includes: (1) self-generated mediators that facilitate the shuttling of electrons from the cells to the anode; (2) electrically conductive pili, capable of long-range electron transfer; and (3) diffusive exchange of electrons between species via soluble electron shuttles such as H₂ (electron-accepting microbes are methanogens) (Table 5.1).

Table 5.1 Electroactive microorganisms (EAMs) in bioelectrochemical systems, BESs

5.2 Hydrogen as the Main Product Using Microbial Electrolysis Cells

Previously, the process of hydrogen generation via electrolyzing dissolved organic matter, using EAMs acting as catalyst, was named “biocatalyzed electrolysis” (Rozendal et al. 2006); subsequently, it was referred to “electrochemically assisted microbial production of hydrogen” (Liu et al. 2005). In earlier research on hydrogen production in MEC, the reactor basically had the similar configuration as the MFC reactor, composed of typical bipolar chamber structure made of glass, where two electrode chambers were isolated by proton exchange membrane (PEM). These initial studies primarily focused on hydrogen generation with acetate as the model compound in MEC. The experimental results showed that the coulomb efficiency (CE, total recovery of electrons from acetate) of MEC could reach up to 60%, which was much higher than that of MFC, reported in the same period. More than 90% of electrons and protons generated via bacterial acetate oxidation were converted to hydrogen. When further considering the maximum conversion rate of hydrogen, assuming 78% of CE and 92% of electron recovery efficiency, it can be easily calculated that 1 mol of acetate can produce approximately 3 mol of hydrogen. Moreover, comparing the hydrogen production capacity of different organic acids (acetate and butyrate), the results showed that acetate was more favorable to the metabolism of microbes in MEC, with hydrogen yield from acetate being higher than butyrate. Preliminary results showed that CE of acetate and butyrate could reach 50–65%, whereas that of glucose only reached 14–21%, which implied the feasibility of exploiting MEC to produce hydrogen and simultaneously degrade end products of dark fermentation (Liu et al. 2005).

$$ {\mathrm{C}}_6{\mathrm{H}}_{12}{\mathrm{O}}_6+{6\mathrm{H}}_2\mathrm{O}\to 12{\mathrm{H}}_2+{6\mathrm{CO}}_2\ \left(\mathrm{thermodynamically}\ \mathrm{unfavorable}\right) $$

(5.6)

$$ {\mathrm{C}}_6{\mathrm{H}}_{12}{\mathrm{O}}_6+{2\mathrm{H}}_2\mathrm{O}\to {4\mathrm{H}}_2+{2\mathrm{C}\mathrm{O}}_2+{2\mathrm{C}}_2{\mathrm{H}}_2{\mathrm{O}}_2 $$

(5.7)

$$ {\mathrm{C}}_6{\mathrm{H}}_{12}{\mathrm{O}}_6\to {2\mathrm{H}}_2+{2\mathrm{CO}}_2+{\mathrm{C}}_4{\mathrm{H}}_8{\mathrm{O}}_2 $$

(5.8)

The stoichiometric yield of hydrogen production from glucose as the substrate would be 12 mol H₂/mol glucose, but this process (Eq. (5.6)) would require a large amount of energy and is unlikely to occur ($\varDelta {G}^{0^{\prime }}$=+3.2 kJ mol). This would be translated into extremely low hydrogen yields when hydrogen is produced from glucose, with acetate and butyrate as the only fermentation by-products. Theoretically, 4 mol H₂/mol glucose can be obtained if only acetate is produced, while only 2 mol H₂/mol glucose when butyrate is the exclusive end product. Not surprisingly, usually only 2–3 mol H₂/mol glucose can be produced in actual fermentation process (albeit some thermophilic strains, e.g., Thermothoga can also reach yields around 3.5 mol H₂/mol glucose) (Logan 2004). In a combined process with glucose fermentation to (2 moles of) acetate and subsequent conversion of acetate in an MEC, the overall hydrogen production yield reached 8–9 mol H₂/mol glucose, while the supplied energy requirement (by external voltage) was equivalent to 1.2 mol H₂/mol glucose (Liu et al. 2005).

In principle, MFC and MEC have similar functional microbes, including bacteria capable of extracellular electron transfer and other collaborative bacteria. Consequently, in the early stage of MEC research, the start-up mode was basically to first adapt the inoculum to obtain the corresponding functional flora by virtue of MFC electricity production, and then shifting into the MEC reactor operation (Cheng and Logan 2007; Call and Logan 2008; Hu et al. 2008; Call et al. 2009; Guo et al. 2010). Thus, in order to obtain an anodic syntrophic consortium between fermentative and anode respiring bacteria (ARB), Montpart et al., for example, first utilized the effluent from an already working MFC, composed of ARB (Montpart et al. 2015). The inoculum was fed with acetate and propionate, and subsequently with sludge from culture flasks, containing fermentative bacteria, in order to develop the syntrophic consortium. Once the syntrophic consortium had colonized well in MFC, the biologically enriched anode was transferred into a single-chamber MEC, treating synthetic wastewater (comprising different complex carbon substrates, i.e., glycerol, milk, and starch) to evaluate hydrogen production (Montpart et al. 2015). In the study by Liu et al., the authors unraveled the effects of different MEC start-up modes on hydrogen production and microbial communities (Liu et al. 2010). Interestingly, the results indicated that the start-up conditions with applied voltages (MEC mode) had a strong influence on the performances of MEC reactors, from the perspective of both CE and COD removal efficiency, and presented larger effect on gas composition, especially on the production of hydrogen. The hydrogen production of the reactor, directly started as MEC, was generally higher than that of the one initially operated in the MFC mode. Microbial community analysis results further demonstrated that microbial communities developed in MECs were well separated from those present under start-up conditions, implying that reactor operation affected microbial community composition (Liu et al. 2010). Subsequently, Lee et al. collected the effluent from an acetate fed-batch MEC operated for over 9 months as an inoculation to the upflow single-chamber MEC, reaching a production rate of 4.3 ± 0.06 m³ m⁻³ d⁻¹ of H₂ with 27–49 kg m⁻³ d⁻¹ removal rate of COD (Lee and Rittmann 2010).

Various pure substrates have been well investigated in two-chamber MECs for hydrogen production (Cheng and Logan 2007), including glucose, cellulose, and various fermentative products (acetate, butyrate, etc.). Near-stoichiometric yields have been obtained by those MEC tests. However, mixed substrates or complex organic matters are still leading to low conversion yields. In summary, simple or pure substrates, like acetate as model substrate, are employed in MECs for mechanism analysis of electron transfer or electron flow calculation, while mixed or complex substrates are commonly studied for scaling up reactors or practical treatment.

5.2.1 Simple Carbon Sources for Hydrogen Production

Acetate, a by-product of dark fermentation of glucose, was typically used as a model substrate to ferment hydrogen in MEC research.

Rozendal et al. first reported biocatalyzed acetate for electrohydrolysis via EAMs, inoculated from the effluent of an electrochemical cell, which was previously acclimatized with sludge from a full-scale upflow anaerobic sludge blanket (UASB) reactor, treating sulfate-rich papermill wastewater for five months (Rozendal et al. 2006). A relatively large double-chamber MEC reactor with a volume of up to 3.3 L, separated by a cation-selective membrane, was operated under an applied voltage of 0.5 V, achieving a hydrogen production rate of 0.02 m³ m⁻³ d⁻¹. The CE and cathodic electron recovery efficiency reached 92% and 57%, respectively. It is worth mentioned that the previous research pointed out the existence of methanogens at the anode, resulting in hydrogen loss, and speculated about the possible impacts, viz., the loss of partial CE and the decreasing numbers of electrons delivered to the anode by competing for consumption of acetate. Unfortunately, this study did not focus on the phenomenon of the electromethanogenesis and did not further analyze microbial communities. In contrast, the authors assumed that the abovementioned consumption was insignificant compared to the H₂ recovery loss at the cathode, because the H₂ generated at the cathode would diffuse to the anodic chamber and be used as electron donor for biocatalysis.

Chae et al. employed a two-chambered MEC, fed with acetate, to elucidate the effects of applied voltages on the hydrogen production. They found that the hydrogen yields generally increased with applied voltages (from 0.1 to 1.0 V), obtaining a maximum H₂ yield of 2.1 mol/mol acetate. Moreover, the higher voltage implied a higher electron loss at the anode, compared to that of the cathode (Chae et al. 2008). Jeremiasse and colleagues obtained the maximum H₂ production rate using acetate and applying 1.0 V (Jeremiasse et al. 2010). In fact, the applied voltage is crucial for hydrogen formation and also significantly affects the H₂ conversion efficiency. Although hydrogen can theoretically be produced at the cathode by applying a circuit voltage greater than 0.14 V (Rozendal et al. 2006; Liu et al. 2005), in reality, higher voltages are required due to the overpotential. In practice, cathodic hydrogen generation can be considered negligible when applying below 0.30 V (Hu et al. 2008; Chae et al. 2008).

Table 5.2 presents a comprehensive overview of MEC performances obtained in different studies, using simple carbon sources.

Table 5.2 Simple substrates used for hydrogen production in the microbial electrolysis cell, MEC

5.2.2 Complex Carbon Sources for Hydrogen Production

Hydrogen can be generated via biocatalytic electrolysis (MEC) with the potential to efficiently convert a variety of dissolved organic matter and refractory wastes from wastes or wastewaters. Even substrates that were previously considered to be unfitting for producing hydrogen, according to the endothermic conversion reactions, can now be valorized by means of MECs.

There is a limited amount of carbohydrates from waste activated sludge (WAS) suitable for utilization by hydrogen-producing microorganisms; thereby, low H₂ yield is typically harvested from the WAS fermentation. Lu et al. obtained H₂ yields of 15.08 ± 1.41 mg-H₂/g-VSS from alkaline-pretreated WAS, which was 2.66-fold of that with raw WAS (5.67 ± 0.61 mg-H₂/g-VSS) in the two-chamber MEC (TMEC). However, more than 13 times higher H₂ production rate was achieved in the single-chamber MEC (SMEC) with alkaline-pretreated WAS, compared to TMEC (Lu et al. 2012b). Besides carbohydrates, there were other substrates (including proteins and their acidification products, such as volatile fatty acids), supporting hydrogen generation in MECs. In addition, it was further confirmed that electrohydrogenesis can react on both the exo-polymeric compounds and the intracellular ones.

Crop castoffs are considered to be a feasible feedstock for dark fermentation to generate hydrogen, thanks to the simple operation and low-energy requirements (Ghimire et al. 2015); however, this process is always associated to the formation of various by-products, mainly volatile fatty acids such as acetate and butyrate (Pan et al. 2010; Xing et al. 2011). Therefore, the integration of dark fermentation with MEC represents an effective way to convert biomass and main fermentation dead-end products into hydrogen (Marone et al. 2017). In order to further enhance hydrogen yield, Li et al. first investigated the effect of pre-adaptation and acclimatization strategies of the MFC anode biofilm grown on diverse substrates and subsequently transferred to the MEC. A maximum H₂ production rate of 4.52 ± 0.13 m³ m⁻³ d⁻¹ under the highest current density of 480 ± 11 A m⁻³ was achieved in a pre-acclimatized anode fed with butyrate (applying 0.8 V), while the one treated with acetate reached 3.56 ± 0.22 m³ m⁻³ d⁻¹ and 346 ± 11 A m⁻³ (Li et al. 2017). Notably, the H₂ yields and removal efficiency of butyrate were substantially higher than in the case of any other substrates (i.e., corn stalk fermentation, ethanol, propionate, or even acetate) (Li et al. 2017).

Table 5.3 presents a comprehensive overview of complex carbon sources that have been used in MEC studies.

Table 5.3 Complex substrates used for hydrogen production in the microbial electrolysis cell, MEC

5.2.3 Hydrogen Loss Evaluation for Microbial Electrolysis Cells

In practice, hydrogen production is boosted in MECs during the initial operation; however, the production of methane is an inevitable consequence for long-term operation of the mixed flora reactor, in most cases. Undesired H₂ sinks, especially by methanogens, have been a serious issue in MEC operations, although H₂ has a low solubility (i.e., 0.0016 g H₂ can be dissolved into 1 kg water at 293 K). In order to inhibit methanogens’ growth, MEC reactors can be put in aerobic conditions for 10 min after each feed cycle, then replenished with fresh medium, and finally flushed with oxygen-free gas to reestablish anaerobic conditions (Selembo et al. 2009b). Except for bioelectrodes exposed to air intermittently (Call and Logan 2008; Call et al. 2009; Lu et al. 2010), there are other strategies adopted to avoid methanogenesis: (1) operation under lower pH (Hu et al. 2008) or lower temperature conditions (Lu et al. 2011), ultraviolet irradiation (Hou et al. 2014a); (2) washout of methanogens by lowering hydraulic retention time (HRT) (Wang et al. 2009); (3) reducing carbonate concentration (Rozendal et al. 2008); and (4) methanogen inhibitor addition (e.g., 2-bromoethanesulfonate) (Chae et al. 2010). Unfortunately, the abovementioned strategies only focus on repressing methanogenesis but overlook other routes of H₂ consumption, including H₂ oxidized by exoelectrogens, or homoacetogenic microorganisms utilizing H₂ and CO₂ to synthesize acetate (2CO₂ + 4H₂ → CH₃COOH + 2H₂O) (Parameswaran et al. 2009). Both paths are commonly defined as hydrogen recycling between the anode and the cathode (Lee et al. 2009), which does not lead to dramatic H₂ loss, but improves the overpotential loss and prolongs duty cycle, eventually resulting in low H₂ recovery (Parameswaran et al. 2011). It seems to be essential to minimize the diffusion of H₂ toward the anode, to rapidly separate H₂ from the MEC reactor (Lee and Rittmann 2010). Instead of conducting top-down inhibition of methanogenesis, Lu et al. employed a novel approach to actively harvest H₂ by extracting it from the reactor, using a gas-permeable hydrophobic membrane and vacuum, leading to 3.32- to 4.29-folds higher H₂ yield than that of the conventional spontaneous release, without CH₄ detection (Lu et al. 2016). But the decreased biofilm growth, accumulation of foulants, and exorbitant cost related to the membrane will be a big challenge in the future.

5.3 Methane as the Main Product in Integrated Anaerobic Systems

Microbial electrolysis system can improve methane production by electrochemical enhancement process (Villano et al. 2011). Traditionally, the planktonic anaerobic bacteria (PAB) and electrochemically active bacteria (EAB) coexist in MEC and disperse in liquid and electrode surface, respectively (Cheng et al. 2009). Methane production partly depends on electron transfer function of PAB and EAB, which are responsible for the carbon dioxide reduction process. In addition, this process can also make use of electrons supplied by current. Hydrogenotrophic methanogens are generally regarded to exploit H₂ as the sole electron donor to reduce CO₂ for methanogenesis. In reality, recent research has clarified that the electron donor source of hydrogenotrophic methanogens is very extensive. There are mainly two ways through which methanogens directly acquire electrons: (1) supply of electrons through electrodes and (2) microorganisms with extracellular electron transport capability (Rotaru et al. 2014a; Fu et al. 2015).

According to the type of substrate utilization, the potential mechanisms of methane formation in a bioelectrochemical system can be divided into two categories: one is through electron transfer, i.e., by means of DIET from the (bio-)cathode, and the other one is through interspecies hydrogen transfer among hydrogen-producing and hydrogen-consuming methanogens. Furthermore, in some cases, acetate and formic acid are formed by means of DIET from electrodes or via acetate- and formate-producing microorganisms (along with H₂ production); hence, acetate and formic acid can be further decarboxylated by acetotrophic methanogens to produce CH₄. Possible paths of methane formation on the (bio-)cathode are shown in Fig. 5.2.

Fig. 5.2

Possible pathways for methane generation in the microbial electrolysis cell, MEC

5.3.1 Microbial Extracellular Electron Transfer Driving Methane Production

Carbon dioxide, methyl compounds, or acetate can be converted into methane in the microbial methanogenesis process. The fundamental pathways are shown in Fig. 5.3. There are two principal ways of obtaining electron donors for acetoclastic or hydrogenotrophic methanogens to generate methane: one is to directly harvest electrons through electrodes and the other is to utilize microorganisms capable of extracellular electron transport to capture electron donors.

Fig. 5.3

Metabolic pathways of methanogens

Methanosaeta is a typical acetoclastic methanogen that exclusively uses acetate for methanogenesis. Morita et al. found that Geobacter was the dominant bacteria in microbial aggregates in cultured anaerobic digestion (AD) reactors, whereas Methanosaeta is the most abundant methanogen, indicating for the first time a possible DIET process with methanogenic wastewater aggregates. Microbial aggregates possess metallic-like conductance similar to the conductive pili of Geobacter sulfurreducens (Morita et al. 2011). Among microbial aggregates formed by the combination of Geobacter metallireducens and Methanosaeta harundinacea, the former can provide electrons to the latter. DIET between Geobacter and Methanosaeta can be used for methane formation (Rotaru et al. 2014b), which changed the viewpoint that the archaea Methanosaeta exclusively uses acetate to produce methane. In addition, metatranscriptomic analysis further revealed that Methanosaeta also has the capacity to reduce carbon dioxide for methane production in AD reactors. The relationship between Geobacter and Methanosaeta is similar to that between fermentation bacteria and syntrophic methanogens, which is based on electron transfer. Kaur et al. found that Geobacter exhibited a clear overwhelming competition for acetate utilization, compared to Methanosaeta in the open circuit (Kaur et al. 2014). Further, Jung et al. demonstrated positive correlation between external resistance and methanogenesis, also showing that the substrate competition among exoelectrogens and methanogens might be influenced by the same external resistance, thus suggesting that the anode potential can regulate the competition between extracellular electron transfer bacteria and methanogens (Jung and Regan 2011).

Methanosarcina belongs to the facultative acetoclastic methanogen, which can utilize a wide variety of substrates, including acetate, methanol, methylamine, and hydrogen. Rotaru et al. found the evidence that Methanosarcina barkeri participated in DIET with Geobacter metallireducens. The study showed that the exploitation of activated carbon particles can replace pili for long-range electron transport, which implied that conductive materials can act as a substitute carrier for pili to perform electron transfer between species (Rotaru et al. 2014a). The close contact is necessary for the implementation of DIET, which may be attributed to the conductivity of pili, whose conductivity is ~5 mS cm⁻¹ (Malvankar et al. 2011). The applied voltage, driving electrons through the external circuit and stimulating methane formation, is comparable to the transmission method through the pili. Nevertheless, on the contrary, the external circuit is not limited by the protein structure and the transmission scale, which can attain long-range electron transfer and display excellent conductivity.

Hydrogen and formic acid at the cathode can also become electron transfer mediators. This is different from DIET achieved by the conductive mediator, and the (bio-)cathode methanation process is in a way more controllable and expandable. On the other hand, the microbial electrosynthesis process based on carbon dioxide reduction at the cathode can also promote the synthesis of chemical substances such as acetate and formic acid (Lee et al. 2017). Altogether, the (bio-)cathode methanogenesis process through DIET prevailingly comprises: (1) hydrogen produced by electrochemical processes that can diffuse into microorganisms to maintain microbial metabolism; (2) hydrogen produced by redox proteins (such as hydrogenase) then used as electron transport mediators; (3) since electrons can pass through the redox transmembrane protein, such as cytochrome c, they can be transferred from the electrodes into the microbes (Kumar et al. 2017). From a macro perspective, we can conclude that: (1) hydrogen evolution reaction (HER) occurs directly at the cathode (or from microorganisms on the electrode surface, E⁰ = −0.41 V) and then hydrogen is absorbed by hydrogenotrophic methanogens and combined with carbonate to form methane (Wang et al. 2009); (2) certain methanogens receive electrons directly from the cathode and combine it with carbonate to generate methane (E⁰ = −0.24 V) (Cheng et al. 2009; Van Eerten-Jansen et al. 2012); (3) homo-acetogens attached to the cathode surface receive electrons from the electrode and synthesize acetate utilizing carbonate (Nevin et al. 2011), and acetotrophic methanogens utilize acetate to generate methane (E⁰ = −0.28 V).

5.3.2 Electricity-Stimulated Anaerobic Methanogenesis Process Using Different Substrates

Conventional methanogenesis primarily depends on the hydrogen or formic acid for interspecies electron transfer in methanogenic environments (Stams and Plugge 2009). It is energetically difficult to complete proton reduction due to negative redox potential of NADH/FADH₂ (NAD⁺/NADH, E⁰ = −0.32 V, FADH/FADH₂, E⁰ = −0.22 V), when hydrogen is used as the electron transport carrier. Therefore, anaerobic microorganisms usually use ferredoxin Fd (Fd_ox/Fd_red, E⁰ = −0.398 V or lower), as a common redox mediator for catalyzing hydrogen evolution reaction (Stams and Plugge 2009). However, some methanogens do not possess cytochromes; thus, Fd and coenzyme F₄₂₀ (F₄₂₀/F₄₂₀H₂, E⁰ = 0.357 V) can help these methanogens, as the most vital hydrogen scavengers, without cytochrome to oxidize hydrogen at extremely low concentrations. Hydrogen is the common product at the cathode of BESs and is also the electron donor for hydrogenotrophic methanogens, as a bridge for converting biohydrogen to biomethane in MEC. Furthermore, hydrogenotrophic methanogens can be highly enriched at the cathode (Lovley 2017; Siegert et al. 2015), and there is also DIET that does not require hydrogen for catalysis (Cheng et al. 2009). Obviously, Methanococcus maripaludis with knocked out hydrogenases was able to directly obtain electrons from the cathode to reduce carbon dioxide into methane (Lohner et al. 2014).

In addition to the diffusion of hydrogen, formic acid can also be used as an electron intermediary to achieve interspecies electron transfer. The discovery of formic acid transfer pathway was due to the fact that the sole hydrogen transfer rate could not match the methane production rate from butyrate degradation in the bioreactor (Thiele and Zeikus 1988). However, only part of the methanogens can utilize formic acid, even though the transfer diffusion rate of formic acid is 100-fold that of hydrogen. Hence, formic acid also becomes an electron loss during the methanogenesis process. Moreover, it is extremely difficult to evaluate the contribution of hydrogen and formic acid to methanogenesis; on the other hand, the electron transfer process in traditional AD relies on two pathways, resulting in restricted possibilities for methane yield enhancement.

MECs, as emerging technologies for anaerobic wastewaters/wastes treatment and energy recovery, can be regarded as a practical integrative step to address some obstacles of AD, such as poor operational stability, low biogas yields, and qualities (Wang et al. 2020a). Importantly, the integrated electricity-stimulated anaerobic system can treat multiple wastes, regulate the establishment of microbial community structures and electron transfer paths, and dramatically improve energy efficiency and overall systems stability. Bo et al., for instance, employed waste activated sludge as substrate in a MEC reactor (at an applied voltage of 1.0 V) for methane generation and obtained 2.3 times higher rates than the conventional AD (Bo et al. 2014). Furthermore, various kinds of biowastes can be employed in such microbial electrolysis integrated anaerobic systems for methane production, including black wastewater (Zamalloa et al. 2013), digested pig slurry (Cerrillo et al. 2016), distillery wastewater (Feng et al. 2017), food waste leachate (Lee et al. 2017), beer wastewater (Sangeetha et al. 2017), food waste (Zhi et al. 2019), etc. Table 5.4 presents a comprehensive overview of different substrates that have been tested so far.

Table 5.4 Different substrates used for methane production in the microbial electrolysis integrated anaerobic systems

5.4 The Energy Efficiency Calculation Formulas Involved in Microbial Electrolysis Cells

The performances of MEC reactors are typically evaluated using various parameters. In this section, we will present the main calculations, which are primarily based on the products associated to hydrogen and methane metabolism.

5.4.1 Essential Parameters Calculation

H₂ or CH₄ production rate (${Y}_{{\mathrm{H}}_2}$, ${Y}_{{\mathrm{CH}}_4}$):

The daily volumetric H₂ or CH₄ production rates are obtained by dividing the produced volumetric H₂ or CH₄ yield by each cycle and normalize it with the effective working liquid volume of the reactor per day.

$$ {Y}_{{\mathrm{H}}_2}=\frac{A_{{V_{\mathrm{H}}}_2}}{t\cdotp {V}_{\mathrm{liquid}}} $$

(5.9)

$$ {Y}_{{\mathrm{CH}}_4}=\frac{A_{{V_{\mathrm{CH}}}_4}}{t\cdotp {V}_{\mathrm{liquid}}} $$

(5.10)

where ${Y}_{{\mathrm{H}}_2}$ and ${Y}_{{\mathrm{CH}}_4}$ are the H₂ or CH₄ production rate (m³ m⁻³ reactor d⁻¹); ${A}_{V_{{\mathrm{H}}_2}}$ and ${A}_{V_{{\mathrm{CH}}_4}}$ are the average H₂ or CH₄ production for each batch (mL/batch); t represents the residence time of each batch (d, day); and V_liquid is the effective working volume of the reactor.

Coulomb Efficiency (CE)

Coulomb efficiency is used to measure the electron recovery efficiency of the microbial anode (Wang et al. 2020b). CE can be used as the metrics for evaluating the performance of the electrodes in BESs, combined with overpotential, which shows the energy loss at the electrodes (Hamelers et al. 2010). Coulombic efficiency can be an indicator to differentiate the involvement of anodic oxidation and acetoclastic methanogenesis in the removal of acetate. Calculations for the Coulombic efficiency (CE) and COD removal efficiency (CRE) can be found in previous studies (Yang et al. 2019).

$$ \mathrm{CE}=\frac{Q}{Q_T}\times 100\% $$

(5.11)

$$ Q= It=\int i\mathrm{dt} $$

(5.12)

$$ {Q}_T=\frac{b\cdotp F\cdotp m}{M}=\frac{b\cdotp F\cdotp \left({\mathrm{COD}}_{\mathrm{in}}-{\mathrm{COD}}_{\mathrm{out}}\right)\cdotp {V}_{\mathrm{liquid}}}{M_{{\mathrm{O}}_2}} $$

(5.13)

where Q is the actual amount of organic matter via anode microbial degradation; Q_T is the theoretical calculated amount of substrate oxidation; I is the current (A or C s⁻¹), which is calculated by Ohm’s law (I = V_voltage/R_ex) from the voltage (V_voltage) drop measured across the resistor (R_ex); t is the time (s); b is the total number of electrons transferred by oxidation (O₂) of 1 mol substrate, b = 4 (acetate); F = 96,485 C mol⁻¹ is the Faraday constant; COD_inf is the initial chemical oxygen demand of substrate, before the reaction (mg L⁻¹); COD_eff is the final chemical oxygen demand of substrate after the reaction (mg L⁻¹); M is the relative molecular mass of the substrate (g mol⁻¹); ${M}_{{\mathrm{O}}_2}=32\ \mathrm{g}\cdotp {\mathrm{mol}}^{-1}$ is the relative molecular mass of O₂.

Current Density (CD)

There are two approaches to express current density: the first is based on the projected electrode (anode or cathode) area (I_A, A m⁻²) and the other is based on the total reactor effective working volume (I_V, A m⁻³) (Wang et al. 2020c).

Electrode overpotential (EO), taking hydrogen generation as an example:

Anode and cathode overpotential are calculated as (E_{eq, anode}, E_{eq, cathode}):

$$ \mathrm{EO}={E}_{\mathrm{meas}}-{E}_{\mathrm{eq}} $$

(5.14)

where E_meas represents the measured anode/cathode potential (V vs. NHE), while E_eq is the equilibrium/theoretical anode/cathode potential (V vs. NHE), using the Nernst equation (based on the acetate).

$$ {\mathrm{CH}}_3{\mathrm{COO}}^{-}+{4\mathrm{H}}_2\mathrm{O}\to {{2\mathrm{HCO}}_3}^{-}+{9\mathrm{H}}^{+}+{8\mathrm{e}}^{-} $$

$$ {E}_{\mathrm{eq},\mathrm{anode}}={E}_{\mathrm{anode}}^{0^{\prime }}-\frac{\mathrm{RT}}{n_{\mathrm{substrate}}F}\ln \left(\frac{\left[{\mathrm{CH}}_3{\mathrm{COO}}^{-}\right]}{{\left[{{\mathrm{H}\mathrm{CO}}_3}^{-}\right]}^2{\left[{\mathrm{H}}^{+}\right]}^9}\right) $$

(5.15)

$$ {2\mathrm{H}}^{+}+{2\mathrm{e}}^{-}\to {\mathrm{H}}_2 $$

$$ {E}_{\mathrm{eq},\kern0.5em \mathrm{cathode}}={E}_{\mathrm{cathode}}^{0^{\prime }}-\frac{\mathrm{RT}}{n_{{\mathrm{H}}_2}F}\ln \left(\frac{{P_{\mathrm{H}}}_2}{{\left[{\mathrm{H}}^{+}\right]}^2}\right) $$

(5.16)

where ${E}_{\mathrm{anode}}^{0^{\prime }}$ = 0.187 V is the equilibrium anode potential at standard conditions; T = 298 K is the absolute temperature (10⁵ Pa) (Logan et al. 2006); n_substrate = 8 is the number of electrons needed to generate H₂ by oxidizing one mol acetate (Table 5.5); R = 8.314 J K⁻¹ mol⁻¹ is the ideal gas law constant, under the standard biological condition, E_{eq, anode} =  − 0.279 V (Logan et al. 2008). ${E}_{\mathrm{cathode}}^{0^{\prime }}$ is the equilibrium cathode potential at standard conditions, assuming [H⁺] = 1 mol L⁻¹, then ${E}_{\mathrm{cathode}}^{0^{\prime }}$= 0 V; ${n}_{{\mathrm{H}}_2}$= 2 is the amount of electrons needed to generate 1 mol H₂ for hydrogen evolution reaction, under the standard biological condition, E_{eq, cathode} =  − 0.414 V (Logan et al. 2008). When at unit partial H₂ pressure and 30 °C (303 K), the cathode overpotential reduces to (Jeremiasse et al. 2010):

$$ {E}_{\mathrm{eq},\kern0.5em \mathrm{cathode}}=-0.060\ \mathrm{pH} $$

(5.17)

Table 5.5 The number of moles of electrons per mole of common substrate (n), based on the half-cell reactions

5.4.2 The Contribution for Production of Hydrogen and Methane in MECs

The source of hydrogen and methane is evaluated by comparing the volume generated by the current or detected by gas chromatograph, that is, the different contributions of H₂ or CH₄ production coming from the current or the substrate (acetate).

The theoretical production of hydrogen generated based on the measured current and substrate consumption (${T}_{V_{{\mathrm{H}}_2-\mathrm{current}}}$, ${T}_{V_{{\mathrm{H}}_2-\mathrm{acetate}}}$) is given by:

$$ {T}_{V_{{\mathrm{H}}_2-\mathrm{current}}}=\frac{\int It}{n_{{\mathrm{H}}_2}\mathrm{F}}{V}_m $$

(5.18)

$$ {T}_{V_{{\mathrm{H}}_2-\mathrm{acetate}}}=\frac{b_{{\mathrm{O}}_2}\Delta \mathrm{COD}{V}_{\mathrm{liquid}}}{2{\mathrm{M}}_{{\mathrm{O}}_2}}{V}_m\ \left(\mathrm{based}\ \mathrm{on}\ \mathrm{the}\ \mathrm{mearsured}\ \mathrm{COD}\ \mathrm{conceration}\right) $$

(5.19)

$$ {T}_{V_{{\mathrm{H}}_2-\mathrm{acetate}}}=\frac{b_{{\mathrm{H}}_2}{V}_{\mathrm{liquid}}\varDelta {C}_{\mathrm{acetate}}}{M_{\mathrm{acetate}}}{V}_m\ \left(\mathrm{based}\ \mathrm{on}\ \mathrm{the}\ \mathrm{mearsured}\ \mathrm{acetate}\ \mathrm{conceration}\right) $$

(5.20)

The theoretical production of methane generated based on the current and substrate consumption (${T}_{V_{{\mathrm{CH}}_4-\mathrm{current}}}$, ${T}_{V_{{\mathrm{CH}}_4-\mathrm{acetate}}}$) is given by:

$$ {T}_{V_{{\mathrm{CH}}_4-\mathrm{current}}}=\frac{\int I\mathrm{dt}}{n_{{\mathrm{CH}}_4}\mathrm{F}}{V}_m $$

(5.21)

$$ {T}_{V_{{\mathrm{CH}}_4-\mathrm{acetate}}}=\frac{\varDelta {C}_{\mathrm{acetate}}{V}_{\mathrm{liquid}}}{M_{\mathrm{acetate}}}{V}_m\ \left(\mathrm{based}\ \mathrm{on}\ \mathrm{the}\ \mathrm{mearsured}\ \mathrm{acetate}\ \mathrm{concentration}\right) $$

(5.22)

where ${T}_{V_{{\mathrm{H}}_2-\mathrm{current}}}$ and ${T}_{V_{{\mathrm{CH}}_4-\mathrm{current}}}$ represent the theoretical production rate of H₂ or CH₄ generated by the current for every batch, after being normalized to the effective working liquid volume of the reactor, per day (m³ m⁻³ reactor d⁻¹); I is the current (A or C s⁻¹), calculated by Ohm’s law (I = V_voltage/R_ex) from the voltage (V_voltage) drop, measured across the resistor (R_ex); t is the time (s); ∫Idt is the coulombs produced by the current, (C); ${n}_{{\mathrm{H}}_2}$= 2 for H₂ and ${n}_{{\mathrm{CH}}_4}$ = 8 for CH₄ are the number of electrons needed to generate 1 mole H₂ (2H⁺ + 2e⁻ → H₂) or CH₄ (CO₂ + 8H⁺ + 8e⁻ → CH₄ + 2H₂O); F = 96,485 C mol⁻¹ is the Faraday constant; ${b}_{{\mathrm{O}}_2}$ is a conversion factor based on the stoichiometric relation between electrons in COD and H₂ gas, equaling to 1 mol H₂ per 16 g O₂; ${b}_{{\mathrm{H}}_2}=4$ is a conversion factor based on the stoichiometric conversion of the amount of one mol acetate consumed to generate the amount of mol equaling to 4 mol H₂ per 1 mol acetate (CH₃COOH + 4H₂O → 2CO₂ + 4H₂); V_m = 22.4 L mol⁻¹ is the gas constant. ${T}_{V_{{\mathrm{H}}_2-\mathrm{acetate}}}$ and ${T}_{V_{{\mathrm{CH}}_4-\mathrm{acetate}}}$ represent the theoretical volume of H₂ or CH₄ generated by the substrate (acetate) for every fed-batch, based on the acetate converted to methane (CH₃COOH → CH₄ + CO₂) (m³ m⁻³ reactor d⁻¹); ΔCOD and ΔC_acetate are the changes in substrate concentration with every fed-batch (mg L⁻¹); V_liquid is the effective working volume of the reactor, mL; M_acetate = 60.05 g mol⁻¹ is the relative molecular mass of the substrate (acetate).

Thus,

$$ \mathrm{CE}=\frac{T_{V_{{\mathrm{H}}_2-\mathrm{current}}}}{T_{V_{{\mathrm{H}}_2-\mathrm{acetate}}}}\ \left(\mathrm{based}\ \mathrm{on}\ \mathrm{the}\ \mathrm{mearsured}\ \mathrm{acetate}\ \mathrm{conceration}\right) $$

(5.23)

5.4.3 The Energy Calculation Involved in MECs

The overall hydrogen recovery (R_H2) is given by:

$$ {R}_{\mathrm{H}2}=\frac{A_{{V_{\mathrm{H}}}_2}}{T_{V_{{\mathrm{H}}_2-\mathrm{acetate}}}}=\frac{n_{{\mathrm{H}}_2}}{\frac{\int It}{nF}}=\frac{2F{n}_{{\mathrm{H}}_2}}{\int It}\ \left(\mathrm{based}\ \mathrm{on}\ \mathrm{the}\ \mathrm{mearsured}\ \mathrm{acetate}\ \mathrm{conceration}\right) $$

(5.24)

Cathodic hydrogen recovery (${R}_{{\mathrm{H}}_2,\mathrm{cat}}$) is given by:

$$ {R}_{{\mathrm{H}}_2,\mathrm{cat}}=\frac{A_{V_{{\mathrm{H}}_2}}}{T_{V_{{\mathrm{H}}_2-\mathrm{current}}}} $$

(5.25)

The overall hydrogen recovery is used to evaluate the ratio of recovered H₂ compared to the maximum potential H₂ recovery, based on the substrate utilization (Wagner et al. 2009). Cathodic hydrogen recovery is used to evaluate the fraction of electrons that form H₂ from the overall amount of electrons reaching the cathode, namely generating current.

Electron reduction efficiency (E_e) is given by:

$$ {E}_e=\frac{Q_{{\mathrm{CH}}_4}}{Q}\times 100\% $$

(5.26)

$$ {Q}_{{\mathrm{CH}}_4}=8\cdotp {n}_{{\mathrm{CH}}_4}\cdotp F $$

(5.27)

Ideal gas law:

$$ {n}_{{\mathrm{CH}}_4}=\frac{PV}{RT} $$

(5.28)

where this equation is used under experimental condition (25 °C, 1 atm, i.e., 1011.325 kPa).

Electron reduction efficiency (E_e) is used to measure the capacity of electron reduction by catalysis at the cathode surface.

The total input energy (W_input) is given by:

$$ {W}_{\mathrm{input}}={W}_{\mathrm{electricity}}+{W}_{\mathrm{substrate}} $$

(5.29)

where W_input is the total amount of energy added to the entire system, kJ; W_electricity (W_E) is the amount of energy added to the circuit by the power source, kJ, adjusted for losses across the resistor; W_substrate (W_S) is the amount of energy added by the substrate (kJ).

The input electricity energy (W_E) is given by:

$$ {W}_E=\frac{\sum_1^n\left(I{E}_{ap}\varDelta t-{I}^2{R}_{\mathrm{ex}}\varDelta t\right)}{1000} $$

(5.30)

where E_ap is the voltage applied, using the power source (V); Δt is the time increment for n data points measured during a batch cycle (s); and R_ex is the external resistor (Ω).

The input substrate energy (W_S) is given by:

$$ {W}_S={n}_S\varDelta {H}_S $$

(5.31)

where n_S represents the substrate consumed (in terms of number of moles) per batch cycle; ΔH_S is the heat of combustion of the substrate, ΔH_acetate = 870.28 kJ mol⁻¹, ΔH_glycerol = 1655.4 kJ mol⁻¹, ΔH_glucose = 2802.7 kJ mol⁻¹ (Selembo et al. 2009b).

The total gained energy (W_gained) is given by:

$$ {W}_{\mathrm{gained}}={W}_{{\mathrm{H}}_2}+{W}_{{\mathrm{CH}}_4}={n}_{{\mathrm{H}}_2}\varDelta {\mathrm{H}}_{{\mathrm{H}}_2}+{n}_{{\mathrm{CH}}_4}\varDelta {\mathrm{H}}_{{\mathrm{CH}}_4} $$

(5.32)

where ${W}_{{\mathrm{H}}_2}$ and ${W}_{{\mathrm{CH}}_4}$ are the energy content generated from H₂ or CH₄ (kJ); ${n}_{{\mathrm{H}}_2}$ and ${n}_{{\mathrm{CH}}_4}$ are the number of moles of H₂ or CH₄ produced during a batch cycle; $\varDelta {\mathrm{H}}_{{\mathrm{H}}_2}$= 285.83 kJ mol⁻¹ and $\varDelta {\mathrm{H}}_{{\mathrm{CH}}_4}$= 890 kJ mol⁻¹ are the calorific values of H₂ and CH₄, based on the heat of combustion (upper heating value).

The methane revenue (${R}_{{\mathrm{CH}}_4}$) is given by:

$$ {R}_{{\mathrm{CH}}_4}={P}_e\frac{Y_{{\mathrm{CH}}_4}\varDelta {\mathrm{H}}_{{\mathrm{CH}}_4}}{V_m}\upeta $$

(5.33)

where P_e = 0.10 £ kW⁻¹ h⁻¹ is the standard price of electricity (referenced from business rates in the UK) (Aiken et al. 2019); ${R}_{{\mathrm{CH}}_4}$ is the revenue from methane (£·m⁻³ reactor day⁻¹); η = 35% is the electrical efficiency with a combustion engine as converter.

5.4.4 The Energy Recovery Efficiency

The total energy recovery efficiency (η_total) is given by:

$$ {\eta}_{\mathrm{total}}={\eta}_E+{\eta}_S $$

(5.34)

where η_total is the ratio of energy output evaluated by produced H₂ or CH₄ to the total energy input composed of electricity input and substrate (acetate) consumption in the entire system; η_E is the ratio of the energy content of H₂ or CH₄ produced to the input electrical energy required; and η_S is the ratio of output energy evaluated by produced H₂ or CH₄ to the input energy from the consumed acetate.

The electrical energy recovery efficiency (η_E) is given by:

$$ {\eta}_E=\frac{W_{{\mathrm{H}}_2}}{W_E}\ \left(\mathrm{based}\ \mathrm{on}\ {\mathrm{H}}_2\ \mathrm{as}\ \mathrm{the}\ \mathrm{main}\ \mathrm{biogas}\right) $$

(5.35)

$$ {\eta}_E=\frac{W_{{\mathrm{CH}}_4}}{W_E}\ \left(\mathrm{based}\ \mathrm{on}\ {\mathrm{CH}}_4\ \mathrm{as}\ \mathrm{the}\ \mathrm{main}\ \mathrm{biogas}\right) $$

(5.36)

The substrate energy recovery efficiency (η_S) is given by:

$$ {\eta}_S=\frac{W_{{\mathrm{H}}_2}}{W_S}\ \left(\mathrm{based}\ \mathrm{on}\ {\mathrm{H}}_2\ \mathrm{as}\ \mathrm{the}\ \mathrm{main}\ \mathrm{biogas}\right) $$

(5.37)

$$ {\eta}_S=\frac{W_{{\mathrm{CH}}_4}}{W_S}\ \left(\mathrm{based}\ \mathrm{on}\ {\mathrm{CH}}_4\ \mathrm{as}\ \mathrm{the}\ \mathrm{main}\ \mathrm{biogas}\right) $$

(5.38)

The conversion efficiency of substrate (η_substrate) is given by:

$$ {\eta}_{\mathrm{substrate}}=\frac{n_{{\mathrm{H}}_2}\frac{{V_{\mathrm{H}}}_2}{V_m}}{n_{\mathrm{substrate}}\frac{C_{\mathrm{substrate}}{V}_{\mathrm{liquid}}}{M}}\times 100\%\left(\mathrm{based}\ \mathrm{on}\ {\mathrm{H}}_2\ \mathrm{as}\ \mathrm{the}\ \mathrm{main}\ \mathrm{biogas}\right) $$

(5.39)

$$ {\eta}_{\mathrm{substrate}}=\frac{n_{{\mathrm{CH}}_4}\frac{{V_{\mathrm{CH}}}_4}{V_m}}{n_{\mathrm{substrate}}\frac{C_{\mathrm{substrate}}{V}_{\mathrm{liquid}}}{M}}\times 100\%\left(\mathrm{based}\ \mathrm{on}\ {\mathrm{CH}}_4\ \mathrm{as}\ \mathrm{the}\ \mathrm{main}\ \mathrm{biogas}\right) $$

(5.40)

where η_substrate is the substrate conversion efficiency; n_substrate is the electron per single mole of substrate; ${n}_{{\mathrm{H}}_2}$ and ${n}_{{\mathrm{CH}}_4}$ are the electrons yielded by H₂ or CH₄; and C_substrate is the substrate concentration (mg L⁻¹).

5.5 Efficiency Improvement for Electron Transport

The electron transfer process is critical for the methanogenesis on the microbe-electrode interface. The electron transfer process of the cathode is similar to that of the anode, except for the direct electron transfer by direct contact and an indirect electron transfer process using hydrogen as a mediator (Miriam et al. 2011). Previous studies have illustrated that hydrogen is an important electron intermediate in the formation of cathodic methane, as well as an important electron donor for basophilic hydrogenotrophic methanogens. Compared with other pathways (Fig. 5.3), hydrogen as an electron donor has the advantage of facilitating the enrichment of a wider range of hydrogenotrophic methanogens by diffusion, which is beneficial to improve the methane production rate. Therefore, it is more advantageous to enhance the hydrogen-to-methane pathway by strengthening the hydrogen evolution reaction for the enrichment of hydrogenotrophic methanogens, viz., promoting the hydrogen-mediated electron transfer process.

The formation of hydrogen at the cathode primarily depends on the electrochemical reaction process. Different electrode materials can affect the electron transfer rate and thus restrict the rate of HER. Furthermore, the characteristics of the biofilm also trigger differences in electron recovery efficiency, which further affect hydrogen yield. On the reaction interface between electrodes and microorganisms, the final electron acceptor is influenced by microorganism types and material properties. Under the conditions of non-pure cultures and non-specific materials, the electrons transmitted to energy metabolism and anabolic processes are different (since diverse microorganisms have different electronic respiratory chains), consequentially resulting in a variety of products. As a consequence, many undesirable by-products are ultimately produced, which affects the electron recovery efficiency.

The microbial community structure plays a decisive role in the distribution of reactive products, and the properties of the electrode materials can cooperate with the microorganisms to capture certain specific electron acceptors, subsequently resulting in high electron transfer recovery. In order to further strengthen bioenergy recovery, the electron transfer process is primarily facilitated in terms of electrode material modification and microbial community regulation. On the one hand, it can promote the rate of electron transfer on the interface of the bioelectrodes, improving the ability of catalyzing HER on the cathode. Furthermore, it can increase the recovery efficiency of electron transfer to the target end products.

5.5.1 Cathode Materials Upgrading

To date, one of the main drawbacks of BESs large-scale application, particularly with MEC, is the demand for costly materials, e.g., platinum in cathode. These materials are often favored due to the dramatic electrocatalytic activity for H₂ evolution, although the performance is negatively influenced by a number of different components that can be found in waste streams. Therefore, more sustainable and low-cost cathodes for bioenergy production via BESs are becoming urgent (Villano et al. 2010). Recently, microbial biocathodes have exhibited more widespread applications, e.g., bioremediation systems for biological reduction of oxidized contaminants (Aulenta et al. 2008, b), biological reduction of nitrates to nitrogen (Clauwaert et al. 2007), or electrochemical reduction of CO₂ to CH₄ (Villano et al. 2010).

In general, upgrading electrode materials primarily focus on reducing the mass transfer resistance of materials and the catalytic resistance. Based on the low hydrogen evolution potential of nickel foam (NF), the high catalysis efficiency of earth-abundant transition metal phosphides, and low cost, Cai et al. studied a one-step phosphorization of NF; the authors used phosphorous vapor to fabricate a 3D biphasic Ni₅P₄-NiP₂ nanosheet matrix, acting as an electron transfer cathodic tunnel for H₂, coupled with a bioanode (Cai et al. 2018). A productivity of 9.78 ± 0.38 mL H₂ d⁻¹ cm⁻² was obtained, which was 1.5-fold higher than NF alone, and even higher than that described for commercially available Pt/C of 5.28 mL d⁻¹ cm⁻² (Cai et al. 2016a) and 4.94 mL d⁻¹ cm⁻² (Hou et al. 2014b). In addition, in order to replace the precious metal Pt, many transition metals, e.g., molybdenum, stainless steel, nickel foam, and other materials, are used as the matrixes, which can be further modified to improve the electrocatalytic activity of the electrode. Selembo et al. compared the effects of different stainless steel alloys (SS 304, 316, 420, A286) and nickel alloys (Ni 201, 400, 625, HX) using sheet metal cathodes in MEC, on hydrogen production, and found SS A286 displayed the best performance of 1.5 m³ H₂ m⁻³ d⁻¹ at 0.9 V (Selembo et al. 2009a). Call et al. confirmed that the stainless steel brush cathode with specific surface area can achieve the maximum productivity of 1.7 ± 0.1 m³ H₂ m⁻³ d⁻¹ at 0.6 V, compared to graphite brush cathode and flat stainless steel cathode (Call et al. 2009). Similarly, Su et al. also found that 3D macroporous stainless steel fiber felt cathode with high electrochemical active surface area has superior catalytic properties for H₂ evolution, achieving 3.66 ± 0.43 m³ H₂ m^-3 d^-1 (current density of 17.29 ± 1.68A m^-2) at 0.9 V (Su et al. 2016). Hrapovic et al. successfully electrodeposited Ni on porous carbon paper, as cathode, obtaining the maximum H₂ production rate of 5.4 m³ m⁻³ d⁻¹, when Ni loaded between 0.2 and 0.4 mg cm⁻², on the contrary, no any increase of hydrogen production under the coelectrodeposition of Pt and Ni (Hrapovic et al. 2010).

Figure 5.4 presents a comprehensive overview of cathode materials that have been used in MEC studies to enhance hydrogen and methane production.

Fig. 5.4

Different cathode types of materials used in the microbial electrolysis cell, MEC

5.5.2 Functional Microbial Community Regulation

In the traditional methanogenesis process, various volatile acids (such as propionate, butyrate, valerate, etc.) need to be converted into acetate and hydrogen by acetogenesis, before being used by acetoclastic and hydrogenotrophic methanogens, respectively. It is generally believed that 70% of the source of methane is derived from acetate and 30% from the contribution of hydrogen (Angenent et al. 2004). However, the growth rate of methanogens is slower than that of fermentative microorganisms, which limits the increase in the rate of methanogenesis. In view of this limitation, a methanogenic process based on extracellular electron transfer is developed inside a conventional anaerobic bioreactor (Liu et al. 2016a). This pathway allows direct electron transfer at the anode, using coenzyme cytochrome c with both oxidized and reduced states, extending the substrate types for methanogenesis. During extracellular electron transport of methanogenesis, the EAB can utilize various types of substrates, such as acetate and propionate, while the electrons generated by oxidizing substrates can pass through an external circuit to drive the reduction of carbon dioxide at the cathode to form methane. Meanwhile, hydrogen also generates from electrons and protons, which can promote the growth of hydrogenotrophic methanogens, counteracting the limitations of traditional acetoclastic methanogens.

Cai et al. coupled the AD with the MEC to treat waste activated sludge and enhance methane generation (Cai et al. 2016b). Based on the results of Illumina MiSeq sequencing, methanogens were enriched in the cathode biofilm (particularly hydrogenotrophic Methanobacterium and acetoclastic Methanosaeta), with two primary methanogenic pathways taking place at the cathode. This implied the possibility of increasing methane production, while reducing WAS digestion time, by controlling the bioelectrochemisty of the process.

Quorum sensing is an essential strategy for microbial community regulation and cell-to-cell communication in biofilms. The principle is that microbes secrete signaling molecules to affect the physiological activities of surrounding microorganisms, such as mobility, sporulation, biofilm formation, virulence, but also symbiosis, competition, toxicity, antagonism, antibiotics production, etc. (Miller and Bassler 2001). Acylated homoserine lactones (AHLs) are a representative signaling molecule that can also be used to regulate the interspecies communication process. AHLs can be synthesized by bacteria like Pseudomonas sp. and degraded via chemical or biological pathways. In recent studies, it has been shown to enhance the respiratory activity associated with electron transfer, facilitating the capacity of electron transfer between cells and electrodes (Toyofuku et al. 2007). This increase in respiratory activity is mainly attributed to two strategies: one is by changing the physiological characteristics of microorganisms, including the cell membrane transmittance (Yong et al. 2013) and gene expression (Hu et al. 2015) (acting directly on the microorganism itself), and the other is by regulating processes related to electron shuttles for the biosynthesis, such as phenazines (Rabaey et al. 2005) (acting on the extracellular synthesis). Both strategies have been demonstrated to effectively increase the electrochemical activity of microorganisms. Cai et al. employed short-chain AHLs (3OC6), as intraspecific signaling molecules to modulate the biofilm community of bioelectrodes in single-chamber MECs. Surprisingly, the overall performance parameters of MECs with AHLs addition were significantly enhanced, including hydrogen yields, CE, electron recovery efficiency, and energy efficiency (Cai et al. 2016c). The lower internal resistance of reactors was verified via electrochemical impedance spectra (EIS). Noticeably, more EAB and fewer hydrogen scavengers, especially homo-acetogens Acetoanaerobium and Acetobacterium, and methanogens, especially hydrogenotrophic methanogen Methanobrevibacter, were detected in cathodic microbial aggregation (Fig. 5.5), which further confirmed the potential of regulating microbial communities by AHLs for strengthening electron transfer and hydrogen production in MEC, and impeding methanogenesis without any chemical inhibitors added (Cai et al. 2016c).

Fig. 5.5

The comparison of specific functional microbial community compositions on genus level between the anode and the cathode in the presence or absence of acylated homoserine lactones, AHLs. (a) is based on three functional microbial categories; (b–d) are subdivided into the specific categories of functional genera classification, including electrochemically active bacteria, EAB, homo-acetogens, and methanogens (reproduced from Cai et al. 2016c)

5.6 Bottlenecks and Challenges

MECs provide a promising potential to boost renewable hydrogen and methane generation from biowastes, possibly providing a new horizon to address imminent challenges in the energy sector, related to the rapidly growing population and fast developing industries. Importantly, MEC as an environmental-friendly technology not only displays a sustainable role in bioenergy recovery, but also simultaneously disposes and valorizes wastes. Undoubtedly, there are big challenges, such as fluctuating performances of MECs and costs of large-scale units, significantly constraining the transfer of bioelectrochemical technology from the laboratory to full scale. For example, system architecture, operating parameters, biological parameters determination, and techno-economic evaluation (such as products revenue, reproducibility, durability, scalability, etc.) need to be implemented and optimized, in order to bring this technology closer to the market.

Noticeably, there are four significant issues that need to be taken into consideration: (1) seeking alternative renewable energy, like solar, wind, waste heat, geothermal and marine energy, to improve sustainability or input energy saving; (2) increasing purities of biogas (H₂ or CH₄); (3) optimizing the electrode space layout to maximize efficiency in the limited reactor space; and (4) catalyzing the surface characteristics of the electrode and evaluating the electrochemical parameters of the composites. In the future, it will be strategic to integrate MECs with other waste treatment technologies to expand their scopes and applications. According to reactor configurations, alloy metal materials (like stainless steel mesh) will allow to form different configurations with large surface area, low overpotential, and low internal resistance of the system, at the same time promoting functional microorganisms adhesion to the electrodes. For the development of the electrode module system, it is necessary to pay more attention not only to the microstructure and the material properties of the electrodes, but also to three-dimensional electrode structure configuration with engineering application potential. The engineering application of MEC technology is promoted by modifying the conductive polymer on a single substrate, applying nanomaterials to improve electrode conductivity and specific surface area, and enhancing electron transfer performance and catalytic activity of bioelectrodes.

In conclusion, new assembly strategies will be explored, based on micro-nano interface of micro-electrode. Also, high-throughput sequencing, stable isotope labeling, and other scientific methods are comprehensively employed to further reveal the function and structure of electrode microbial flora in depth, and shed light on microbial interactions, as well as extracellular electron transfer mechanism, based on electron mediator, nanowire, and cytochrome. Overall, these approaches are expected to provide technical and theoretical understanding for the development of viable and sustainable applications of MEC technology.