In this chapter, we focus on microbial fuel cells (MFCs) that convert the energy from organic matters into electrical energy using microorganisms. MFCs are greatly expected to be used as a relatively low-cost and safe device for generating renewable energy using waste biomass as a raw material. At present, however, it has not reached the desired practical application because of the low-power generation; hence, improvements on fuel cell efficiency, such as electrode materials, are still being examined. Here, we focus on the microorganisms that can be used as catalysts and play a central role in improving the efficiency of the fuel cells. Several kinds of microbial catalysts are used in MFCs. For example, Shewanella oneidensis has been well studied, and as known, since S. oneidensis transports the electrons generated within the cell to the surface layer, it does not require a mediator to pass the electrons from the cells to the electrode. Furthermore, Escherichia coli and Saccharomyces cerevisiae, a model organism for MFCs, are also used. The improvements of such microbial catalysts have also been proceeding actively. Here, we elaborated on the principle of MFCs as well as the current situation and latest research on the catalyst development.
- microbial fuel cell
- renewable energy
- bioelectrochemical device
The fossil fuel depletion and inevitable global warming have become worldwide problems; thus, significant efforts have been made to generate and utilize renewable energy to alleviate these crises. Methods for obtaining energy compounds from biomass, such as ethanol, methane, and hydrogen, have been developed using environment-friendly technology, and some of these technologies have been put to practical use. It is important to establish the technologies that are able to obtain energy in various forms according to the environments and circumstances of each region. Apart from the above technologies, biofuel cells utilizing microorganisms and enzymes, which can generate renewable electrical energy from organic matters contained in biomass, begin to attract attention as a means to obtain sustainable energy. It has not been put into practical use yet, but without the problem of by-products, electricity can be directly obtained from the devices, whereby multiple operations, such as product distillation (e.g., ethanol), are not necessary. Moreover, if biomass waste is used as the fuel, no food competition will occur. Therefore, using this method, energy can be obtained sustainably (Figure 1).
There are various types of biomass, e.g., sustainably harvested wood, waste paper, food waste, sewage sludge, and various wastewaters. Taking wood-based biomass as a fuel example, when everything is burnt using available technology for thermal power generation, there will be nothing left, and we will lose some other useful compounds contained in it. On the other hand, in biofuel cells, although electricity is generated from the sugar obtained from the biomass, other components in the wood, such as lignin, can be used for purposes other than power generation. Generally, the energy density of the biomass used as a fuel for MFCs is high. For example, glucose and xylose, found in various plant biomass, can produce up to 20 or 24 electrons per molecule, provided that they are completely oxidized to carbon dioxide. It is possible to generate 4430 Wh power per kg of glucose according to the calculation described later. For reference, a typical lithium-ion battery has a weight energy density of about 200 Wh per kg. This comparison means that glucose and xylose are two biofuel sources of interest, especially as electron donors. Therefore, MFCs using glucose and/or xylose as their fuel have great potential as a means of obtaining high energy.
In biofuel cells, biological reactions are used for the oxidation reaction of biomass, and they are divided into two based on the type of catalyst used: (1) enzymes and (2) microorganisms. When enzymes are used, the most widely studied mechanism is the two-electron oxidation system by glucose oxidase (GOx) or glucose dehydrogenase (GDH) . Since purified enzymes are generally used, the reaction rate is faster than using microorganisms. However, the number of electrons obtained by one enzymatic reaction is smaller than that of a microorganism. For example, when GDH is used as the catalyst, glucose is oxidized to gluconic acid, and at most, only two electrons are obtained from one glucose molecule. Therefore, if only one enzyme is used, high-energy production per glucose consumed cannot be much expected. Further, the addition of cofactors, such as nicotinamide adenine dinucleotide and pyrroloquinoline quinone (PQQ), is necessary for the enzymatic reaction to enhance the energy production. Furthermore, the cost of enzyme purification is also high; hence, enzymes are better utilized in sensor applications than energy production. By contrast, as mentioned above, one completely oxidized glucose molecule gives 24 electrons when using microorganisms as the catalyst. It shows a possibility of obtaining more electrons per glucose consumed. Moreover, the addition of cofactors is not necessary, unlike when enzymes are used. These are some substantial advantages of MFCs. However, the production of low power in MFCs is still a problem because of typical processes in living organisms, such as the uptake of glucose into cells, metabolism repression, and extraction of electrons from the inside of cells (Figure 2). Many researchers are working to solve such problems, and those results are summarized in recent review articles [2, 3].
Looking back at the historical background, research on the MFCs has been conducted for a long time, whereby the first idea of using microorganisms to produce electricity was conceived and reported by Potter in 1911 .
The performance of MFCs is evaluated based on some indicators. The electrical energy (Wh) used to express the capacity of dry batteries is also an important indicator, but only a few papers have reported it so far. In most cases, the performance is expressed as the maximum power per anode electrode area (power density per area) or the maximum power per cell volume (power density per volume). The latter is a straightforward index and important for practical use. For example, a relatively high-performance small-scale fuel cell (2.5 mL) using a complex (mixed) microorganism system was reported in 2007 with a power density of 1550 W/m3 . Other fuel cells performing beyond 500 W/m3 were also reported [17, 18, 19, 20, 21, 22], but many of them are still at a microliter or milliliter scale. Owing to the low proton diffusion speed and high internal resistance, the maximum power per volume tends to be small for a large-scale fuel cell. MFCs with a volume more than 1 L were also being studied in the laboratory, but the maximum power per volume was still low at the level of several W/m3 to tens of W/m3 [23, 24]. Scaling-up is also another issue of MFCs, and further improvements are still being conducted.
Practical applications of MFCs are still problematic because of the high cost and low-power generation. Despite this situation, research on the implementation of MFCs has been carried out. For example, an artificial stomach called Gastrobot (aka Chew-Chew train) using
Here, we will explain the mechanism of electron generation in microorganisms, introduce the principle of MFCs, describe the microbial catalysts used for various MFCs mentioned above, and discuss the recent topics on microbial catalysts.
2. General principles of MFCs and various microbial catalysts
2.1. Mechanism of electron generation in microbial cells
MFCs utilize the decomposition energy of organic matters by the organisms to produce ATP, known as the energy currency, based on the energy obtained from this process. Taking glucose decomposition in
2.2. Calculation of the energy obtained from glucose
In the case of using glucose as the fuel source, the reaction occurring in the anode tank is represented by Eq. (1), and the reduction reaction occurring in the cathode tank is represented by Eq. (2).
The oxidation-reduction potential of Eq. (1) is −0.42 V, whereas the oxidation-reduction potential of Eq. (2) is 0.82 V. Therefore, the total potential difference of the MFC reaction (Eq. (1) + Eq. (2)) as represented by Eq. (3) is 1.24 V. Theoretically, the voltage exceeds 1 V, but in most cases, it has never reached that value.
Assuming that 24 electrons are obtained from 1 glucose molecule and that they can be recovered in 1 h, the quantity of electricity (Ah) obtained from the glucose (1 kg) can be calculated using the Faraday constant (96,485 C/mol) as shown in Eq. 4. As a result, the electrical energy of 4430 Wh can be achieved if the potential is 1.24 V; accordingly, this value is the same as the value mentioned in the Introduction section.
2.3. Basic components of dual-chambered MFCs using a mediator
A dual-chambered fuel cell consisting of an anode tank and a cathode tank is the simplest and has been used for a long time for MFCs. In many cases, they are separated by a cation exchange membrane (CEM) to create a potential difference between the two tanks (Figure 4). CEM prevents mixing of each content and allows the protons generated in the anode to migrate to the cathode. In addition, CEM selection, especially based on its proton transfer efficiency, is important because it significantly regulates the movement of the protons responsible for the pH reduction at the anode affecting the activity of microorganisms and the delivery of electrons to the oxygen at the cathode. Also, some factors to consider, such as durability and cost, are important for selecting CEM. At present, Nafion is popular for many MFCs [30, 31].
Numerous research studies are being conducted to evaluate the influence of the electrode materials on the performance and cost of the MFCs. Carbon materials, which are noncorrosive, have been widely used because of their high electrical conductivity and chemical stability, e.g., carbon rod, carbon fiber, carbon felt, and carbon cloth . Biocompatibility, specific surface area, electrical conductivity, and cost are important factors for its selection. Since its discovery in 2004, graphene has been attracting much attention for its use as an electrode because of its high specific surface area, electrical conductivity, and biocompatibility . In fact, graphene has been already used in lithium-ion batteries, and the development of graphene-modified materials to increase the power density has progressed actively [33, 34]. Moreover, since biofilm formation by microorganisms on the electrodes affects the performance of MFCs, the preference of electrode materials tends to shift from two-dimensional to three-dimensional surfaces, where a larger surface area is obtained; thus, the contact with microorganisms increases. Furthermore, metals are also used as the electrodes. The conductivities are higher than those of carbon materials, but they are prone to corrosion in the anode solution. Therefore, metals are problematic to use, except for stainless and titanium. To improve such problem, materials in which metal is incorporated into graphite have been made .
A phosphate buffer or bicarbonate buffer solution is often used for the anode electrode solution to achieve high performance [16, 35]. The pH of the solution affects not only the activity of microorganisms but also the transfer of hydrogen ions used from the anode to the cathode when the electrons are transferred to oxygen at the cathode. The solutions contain microorganisms as the catalyst, organic matter as the fuel, and mediator as the electron carrier. In addition, there are reports that the performance of MFCs was improved by adding NaCl to increase the ionic strength .
Regarding the fuel, many substrates have already been studied . For example, acetic acid, lactic acid, glycerol, glucose, xylose, sucrose, starch, yeast extract, malt extract, various real wastewaters, and synthetic wastewater were used depending on the purpose of each research. Generally, the fermentable substrate of microorganisms is used to generate electricity more efficiently. There is a trend where glucose is used when using
Regarding the mediator, although some microorganisms can carry electrons directly to the electrode as described later, in many cases, the electrons cannot be carried, or the performance is low even if carried, so an artificial mediator that can pass through the cell membrane is added to the anode solution. The typical compounds for artificial mediators are methylene blue, neutral red, 2-hydroxy-1,4-naphthoquinone (HNQ), thionine, benzyl viologen, 2,6-dichlorophenolindophenol, and various phenazines . It was reported that the hydrogenase donates electrons to the neutral red , but the process was not yet clearly proven as to how these mediators deprive electrons of the cell. It is thought that, depending on the type of mediator, the electrons may be taken directly from NADH or obtained from the electron transfer system of the cell membrane. On the other hand, there is also a difficult aspect of using a mediator. In order to increase the electron transfer efficiency by the mediator, it requires a high concentration, but because of its high toxicity, it has a strong influence on the cells; therefore, the level of use is necessary to be controlled.
Finally, the cathode solution is explained as follows. The electrons generated at the anode are carried to the cathode, where the reduction reaction takes place. When oxygen, the most common electron acceptor, is used as an oxidizing agent, aeration is necessary because oxygen has low solubility (about 8 mg/L DO). There are cases where oxygen generation by the photosynthesis of algae is used for oxygen supply [40, 41]. In the reaction at the cathode, H2O is produced by oxygen, whereby the electrons were carried from the anode via an external circuit and protons were carried from the anode solution via CEM. There is also a report that the addition of hydrogen peroxide leads to an improvement in power generation . Besides oxygen, there are various electron acceptors ; for example, an oxidizing agent such as iron ferricyanide is also used for the cathode. In many cases, the ferricyanide has a high mass transfer efficiency and a high cathode potential so that a high output can be obtained. The combination of carbon electrodes and ferricyanides to achieve power 50–80% higher than the combination of Pt/carbon electrodes and oxygen was reported . In the case of using ferricyanide, once the trivalent iron ion receives the electrons, it becomes divalent, and when it delivers the electrons to oxygen, it reverts to the trivalent state. However, the latter reaction is less likely to occur owing to the low solubility of oxygen. Ferricyanide is an excellent electron acceptor, but owing to its toxicity, its use is generally limited to the laboratory. Other than oxygen and ferricyanide, there are also many candidates, for example, nitrate, persulfate, permanganate, and manganese dioxide. It is also possible to use the nitrate contained in the wastewater because its redox potential is close to that of oxygen, and then, the nitrate is reduced to nitrogen gas at the cathode .
2.4. Other types of MFCs
MFCs are typically divided into a dual-chambered cell described above and a single-chambered cell (Figure 5). In the latter, a membrane-type positive electrode with oxygen permeability called an air cathode is used . The electrode is coated with the platinum catalyst, and H2O is produced from the oxygen permeated from the atmosphere, the proton in solution, and the electron from the anode. However, if the permeated excess oxygen diffuses and is delivered to the microorganisms at the anode, the electrons generated in the microorganisms are then transferred to the oxygen, and the energy recovery rate decreases. Therefore, a CEM is used between the anode solution and the cathode in order to prevent a decrease in energy recovery. This single-chambered type has already been widely used currently.
Moreover, there is a mediator-free type that does not require an artificial electron compound . The microbial strains used here can synthesize mediators themselves and/or have an electron transfer function on the cell surface. In the former, the self-synthesized mediators are flavin compounds, hydroquinone, and phenazine that are able to transfer the electrons to the electrode. In the latter, membrane-bound proteins such as pili, c-type cytochromes, and filaments are known as cell surface structures that can directly transmit electrons. The biofilm formation on the electrodes, namely, biocompatibility of the electrodes, is also important for power generation via such direct electron transfer. Therefore, research on electrodes promoting the formation of the biofilm is actively being conducted.
In addition, with the use of MFCs in wastewater treatment, contamination on the CEM results in reduced power generation; hence, membrane-free MFCs have also been studied .
2.5. Various microbial catalysts
Various microorganisms have been studied for a long time since the first experiments on
In such a research situation, there are relatively many examples of research on
3. Recent topics of microbial catalyst and future directions
3.1. Modification of microorganisms
The utilization of chemical and biotechnological techniques is important to modify the microbial cells as biocatalysts in the MFC system. Molecular biology approaches are effective tools to improve the performance of the biocatalysts for the desired system. In this section, recent topics about the approaches for microbial catalyst development are discussed.
3.1.1. Chemical treatment
Mediators and macromolecular catabolic enzymes, which are used for electron transfer and other metabolic activities, are abundant in the cytoplasm of the microbial cells used in the MFCs. However, it is not easy to transport the mediator molecules to the bacterial outer membrane so as to reach the electrode. The lipopolysaccharide (LPS) layer on the Gram-negative outer membrane is compact and nonconductive; thus, most microbial cells are nonconductive . It was found that chemically perforated pores and channels on the cell membranes accelerated electron transfer, leading to an improved power output for an MFC using
3.1.2. Biosurfactant production by gene modification
To increase the cell permeability of biocatalysts in the MFCs, Zheng et al. proposed a new approach by inducing the biosurfactant production based on a genetic modification . It is true that the efficiency of membrane permeability can be improved with a biosurfactant, which ultimately increases the transport across the membrane. In addition, overexpression of the
3.1.3. Introducing the extracellular electron transfer pathway
The sparse availability of genetic tools in manipulating electricity-generating bacteria and the multiple overlapping pathways for extracellular electron transfer make it challenging to modulate electron transfer and/or introduce other functions of interest. In response to this challenge, several studies have taken the complementary approach of engineering portions of the extracellular electron transfer pathways into the well-studied industrial microbe
3.2. Extremophilic microorganisms
An extremophilic microorganism thrives in physically or geochemically extreme conditions that are detrimental to most life on Earth. They thrive in extreme hot niches, ice, and salt solutions, as well as acid and alkaline conditions; some may grow in toxic waste, organic solvents, heavy metals, or several other habitats that were previously considered inhospitable for life. Extremophiles can be used to oxidize sulfur compounds in acidic pH to remediate wastewaters and generate electrical energy from marine sediment microbial fuel cells at low temperatures. The MFC performance of these extremophilic microorganisms has been well summarized in several review papers [49, 64]. In this section, the recent advances of MFCs using extremophilic microorganisms as catalysts are briefly introduced and discussed.
3.2.1. Acidophiles and alkaliphiles
An increase of cell voltage is seen at increasing anode pH because of the additional pH gradient representing a source of energy. The practical implication of an elevated cell voltage is that more energy can be gained from MFC systems at higher pH values. By contrast, operating the anode of MFCs at an acidic condition has an advantage that the protons will not cause diffusion limitations in the cathode compartment for the reduction of oxygen, and therefore, it will not limit the current production . However, under a low-pH condition, the microbial cells have to maintain a near-neutral cytoplasm  which consumes a portion of the energy derived from the electron transport for other processes, such as proton export, that increases the anode overpotential, leading to decreasing power generation. At pH 2.5,
Meanwhile, at high pH, the lower anode potential results in an increased cell voltage. In general, the anode becomes acidified during the MFC operation, and the cathode becomes more alkaline, followed by a reduced cell voltage and power output.
3.2.2. Psychrophiles and thermophiles
Temperature also has a major impact on the output of MFCs. It influences the activity of microorganisms, the electrochemical reactions, and the Gibbs free energy change of the reactions. There exists an optimum temperature for enzymes in the MFCs, and the electrochemical reaction rate increases with increased temperatures. A lower operating temperature adversely influences the output, start-up time, and substrate oxidation rate in the MFC system. This negatively impacts the MFCs for processes such as wastewater treatment because the streams are generally at low temperatures. However, the advantage of low temperature for the MFCs is that they typically produce higher Coulombic efficiencies [72, 73]. The microbial community was enriched from the anaerobic sludge at the anode of an acetate-fed MFC operated at 15°C with psychrophiles
On the other hand, the advantages of operating at high temperatures are higher microbial activity, better substrate solubility, high mass transfer rate, and lower contamination risk. An example of an improved current generation at a high temperature (60°C) is a marine sediment MFC that generated 209–254 mA/m2 compared with 10–22 mA/m2 at 22°C . Recently, an MFC with a higher operating temperature (70°C) has generated 6800 mA/m2 . Furthermore, the hyperthermophilic MFCs were operated at above 80°C . However, a negative point of thermophilic MFCs is higher rates of evaporation than the MFC system itself. Therefore, a continuous mode of MFCs was proposed to allow a replacement of the anolyte and catholyte .
In this section, recent topics of microbial catalysts for MFCs were introduced. There are two approaches in developing the microbial catalysts. One is the modification of existing microorganisms using chemical treatments or biotechnological techniques, including gene editing. The other is exploring new microorganisms from the environment, including extreme conditions. Although new findings and knowledge were obtained from both approaches, a drastic improvement on the MFC performance to achieve a paradigm shift has not appeared yet. In parallel with the improvement of microbial catalysts, the development of the fuel cell system, including the electrodes, was intensively studied to increase the output of MFCs. In particular, the application of graphene-modified electrodes  and the investigation of electron acceptors  have shown remarkable progress in the past decade. In order to actualize the practical use of MFCs, a synergistic impact from the combination of microbial catalyst and fuel cell system is essential.
This chapter focuses on the significance of MFC development, the historical background and fundamental principles of MFCs, and their recent developments, especially on microbial catalysts. MFCs have not reached the desirable level of power generation that supports daily life because of the problems such as scaling-up. On the other hand, developments of technology combining wastewater treatment and power generation, and application for environmental sensors are progressing to a stage close to practical use. If these popularizations continue, it will further develop its application in broader fields. Owing to their limitations, it may be difficult to force MFCs to become the main power supply in our daily life in the future, but it seems possible to use them as an auxiliary power supply. In addition, MFCs may become useful as a power supply in areas where the infrastructure is not well developed, for example, a portable power supply generating electricity if water is added. Regarding microbial catalysts, it is also known that various microorganisms can generate electricity, and if this superior power-generating function of these microorganisms can be integrated into a microbial cell using the synthetic biological method developed recently, the ability of the microbial catalyst will dramatically increase. Soon, its power generation ability could be greatly improved in combination with the progress of other constituents.
This work was supported by JSPS KAKENHI Grant Number JP17K06932.
Conflict of interest
The authors declare no conflicts of interest associated with this text.