Towards effective small scale microbial fuel cells for energy generation from urine

To resolve an increasing global demand in energy, a source of sustainable and environmentally friendly energy is needed. Microbial fuel cells (MFC) hold great potential as a sustainable and green bioenergy conversion technology that uses waste as the feedstock. This work pursues the development of an effective small-scale MFC for energy generation from urine. An innovative air-cathode miniature MFC was developed, and the effect of electrode length was investigated. Two different biomass derived catalysts were also studied. Doubling the electrode length resulted in the power density increasing by one order of magnitude (from 0.053 to 0.580 W m -3 ). When three devices were electrically connected in parallel, the power output was over 10 times higher compared to individual units. The use of biomass-derived oxygen reduction reaction catalysts at the cathode increased the power density generated by the MFC up to 1.95 W m -3 , thus demonstrating the value of sustainable catalysts for cathodic reactions in MFCs.


Graphical Abstract 1 Introduction
In the face of the growing problem of fossil fuel depletion, there is global interest in developing sustainable and environmentally friendly forms of energy. One form of alternative energy that may be viable in addressing this problem is bioenergy [1,2]. In this context, Microbial fuel cells (MFC) hold great potential as green and carbon-neutral technology that directly converts biomass into electricity [3].
MFCs are electrochemical devices that take advantage of the metabolic processes of microorganisms to directly convert organic matter into electricity with high efficiencies for long periods of time [4]. Compared to other bioenergy conversion processes (i.e. anaerobic digestion, gasification, fermentation), MFCs have the advantage of reduced amounts of sludge production [5], as well as cost-effective operation, since they operate under ambient environmental conditions (temperature, pressure) [6]. Moreover, MFCs require no energy input for aeration so long as the cathode is passively aerated, for example via the use of a single-chamber device [7]. Lastly, MFCs have the ability to generate energy remotely by using a range of feed stocks, and can thus be used in areas of poor energy infrastructure.
Organic waste used as a feed stock in particular offers attractive prospects from its costeffectiveness and abundance. Urine has been demonstrated to be an effective feed stock for MFC operation with the additional benefit of nitrogen, phosphate and potassium recovery from the fuel [8]. In particular, according to Ieropoulos et al [9], urea is enzymatically hydrolysed to ammonia and carbon dioxide. Ammonia is then oxidised at the anode of the MFC to generate mainly nitrite and in smaller amounts nitrate [10].
Despite the breadth of applications and the growing interest in MFC technology over the past two decades, commercialisation of MFCs for energy generation has not yet been realised.
The major limiting factors that hinder the practical implementation of MFCs at large scale, are the cost of materials used and the difficulties in the scale-up process [11].
Typically the electrodes are made from highly cost-effective materials such as carbon cloth, carbon paper, and graphite based rods, plates and granules. Recently, even some metals, such as copper and silver, have been shown to be effective anode materials [12]. However, expensive metals, such as platinum, are usually used at the cathode to enhance the oxygen reduction reaction (ORR) [13][14][15]. Recently, the use of biomass-derived catalysts recovered from waste has been proposed as an effective alternative to expensive metal ORR catalysts. In particular, biomass-derived materials from wood [16], sewage sludge [17] and bananas [18] have been shown to function as ORR catalysts to boost MFC performance whilst reducing the device cost and its carbon footprint. Doping these materials with heteroatoms such as nitrogen and sulphur [19], also in combination with nanoparticles like iron [20], has been shown to enhance the catalytic activity towards the ORR even further.
Another limitation towards practical implementations of MFCs, is their poor performance due to high internal resistances and ohmic losses experienced upon scale-up [21]. Consequently, the power performance of MFCs is low compared to other renewable energy technologies [8,22] This approach has been referred as the 'miniaturisation and multiplication' strategy [9].
MFC miniaturisation offers other advantages as well. The large surface area-to-volume ratio and short electrode distances -typical characteristics of miniature MFCs-provide a pathway to reducing ohmic losses, improving the mass transport processes between bulk liquid, biofilm and electrode and therefore enhancing power performance [23]. The consolidation of microfabrication techniques has led to the first prototypes of micro-sized MFCs, which have been discussed in a recent review [11]. Nonetheless, the process of miniaturisation of the MFC technology is still in its infancy. The two-chamber configuration is typically adopted for the miniature MFCs reported thus far, and, usually, a ferricyanide solution is used as the catholyte [24]. Given the greater operational simplicity and cost-effectiveness of oxygen diffusion systems, air-cathode MFC designs should be considered instead. Moreover, a more in-depth analysis on how to effectively miniaturise the system for better performance would be beneficial.
With the aim of guiding the development of efficient small-scale MFCs, this study reports the development of an innovative air-cathode small-scale MFC and analyses the effect that the chamber length (and therefore the electrodes length) have on its performance either when operated as a single unit or when assembled in a stack. No expensive metals have been employed at the cathode, and the use of two types of innovative and highly sustainable biomass-derived ORR catalysts are compared with a catalyst-free device.

Materials
All reagents used were of analytical grade and purchased from Sigma-Aldrich and Alfa Aesar.
Unless otherwise stated, all aqueous solutions used were prepared with reverse osmosis purified water. Polydimethylsiloxane (PDMS, Dow Corning Sylgard 184) was purchased from Ellsworth Adhesives (UK).
Artificial Urine Medium (AUM) was used as the feedstock and prepared as previously described [25]. Tetrasodium pyrophosphate was added to the AUM as a precipitation inhibitor. The resulting feedstock was sterilised by filtration (Grade p8 filter paper, Fisher Scientific, UK) prior to use.

Microbial Fuel Cells
Two geometries were used in this study, leading to the fuel cells MFC_S (for short length) The proton exchange membrane (Nafion® 115, Sigma-Aldrich) was hot pressed to the cathode by applying a pressure of approximately 2.5 bar for 12 minutes at a temperature of 150 °C.

Use of a biomass-derived oxygen reduction reaction catalyst
Two different biomass-derived ORR catalysts, named as BC1 and BC2, produced by hydrothermal carbonisation, were tested at the cathode of MFC_L. Both catalysts were synthesised from glucose and ovalbumin as described in [26] and [19]. BC1 is a nitrogen doped carbon aerogel, while BC2 is a nitrogen and sulphur co-doped aerogel that was prepared with an additional iron source. A loading of 1.5 mg per cm 2 of the cathode area was used for each ORR catalyst. 1.5 mg of catalyst was mixed with 105 μL of Nafion® perfluorinated resin solution and sonicated for 3 minutes. The resulting suspension was spread over 1 cm 2 of carbon cloth. Once dried, the doped cathode was bound to the Nafion® membrane as shown in Figure 1 above. The MFCs with the doped cathodes were named as MFC_BC1 and MFC_BC2, according to the ORR catalyst used.
The morphology of the resulting electrodes was characterised using a Hitachi S-4300 scanning electron microscope (SEM).

Operation of the MFCs
All MFCs were fed with AUM at the flow rate of 0.36 mL min -1 (hydraulic residence times of The internal resistance (R int ) of the MFC was calculated from the linear fit of the ohmic region of each polarisation cell potential curve (R int = ΔV/ΔI), as previously described [3].

Stacking
To scale-up the power output, MFC units with the same geometry were electrically stacked in series and in parallel, as shown in Figure 2. The MFCs were enriched individually and stacked after the five days of enrichment, once a steady current was generated. Once stacked, the MFC units were fed in parallel with AUM and no bacteria. The polarisation experiments on the stack were performed after at least 24 hours of operation.

Calculations
The maximum current density (I max ) under mass transport limiting conditions at the electrode, is expressed according to [27] as: Where n is number of electrons equivalent corresponding to the limiting compound The Reynold's number (Re) and mass transfer coefficient (k C ) for laminar flow in a channel is defined as [28]: : Where H is the height (m), and L is the lateral dimension length (m).
The diffusion-layer thickness (λ) at the electrode surface was calculated with the following equation:

Effect of Electrode Length on Performance
The influence of the electrode length on the performance of small scale MFCs, was The increase in power and current density is suspected to be due to an increase in the mass transfer between the bulk fluid, biofilm and electrode surface. When observing the cross section of a MFC square electrode chamber, the height, H, and the lateral dimension (length), L, will affect the performance of the device. On one hand, when the height of the channel is reduced (i.e. the distance between electrodes is reduced) in the MFC, the miniaturised device benefits from a greater rate of mass transfer due to an increase in the surface area to volume ratio of the device [23]. As a result, the power density generated by miniature MFCs is higher than large-scale devices [32]. On the other hand, when the lateral dimension of the channel (length), L, of the electrode chamber is increased, the hydraulic diameter of the channel is increased as per Equation 4. Consequently, the mass transfer coefficient, k C , will increase as per Equations 2 and 3. Therefore, when L is increased, whilst maintaining a fixed H, the mass transfer coefficient is increased, and hence the diffusion-layer thickness at the electrode surface will decrease (Equation 5). By altering the length of the channel, the maximum current density available at the electrode will therefore increase (Equation 1), and, consequently, result in high fuel consumption efficiency and an improvement in power performance. Figure 4 demonstrates that increasing the length of the flow channel, for a fixed flow rate, will increase the mass transfer coefficient and decrease the diffusion-layer thickness. Values here have been calculated using Equations 1-5, with the flow rate at 0.36 mL min -1 , and a linear velocity of 22.5 mm min -1 . For urine, the kinematic viscosity (μ / ρ) is estimated to be 1.07 mm 2 s -1 at 20 °C [33], and the diffusivity of urea in water is 0.082 mm 2 min -1 [34].
To ensure that these assumptions are valid, the flow regime in the flow channel must be laminar. This is confirmed by the Re values for MFC_S and MFC_L, which are 1.4 and 1.9 respectively, as calculated by considering L values of 4 and 8 mm, for MFC_S and MFC_L respectively, and H equal to 4 mm.
By increasing the length of the electrode in the MFC devices, a better fuel efficiency has been achieved, with consequent improvement in performance [35]. This is in accordance with a recent study by [36] whereby increasing the length of a graphite fibre brush anode from 12 mm to 30 mm the power density increased from 1.13 to 1.65 W m -2 . The better supply of redox species (c) to the anode leads to an increase in the measured current density (I), according to equation 6: Where: n is the moles of electrons involved in the reaction; F (C mol -1 ) is the Faraday constant; K c (m s -1 ) is the mass transfer coefficient; λ (m) is the diffusion layer thickness; c is the concentration of the redox compound (mol m -3 ).

Stacking the Miniature MFCs
To scale up the power output, MFC_S and MFC_L were arranged in stacks of three units each. The MFC_S units were electrically connected either in parallel or in series to evaluate the best configuration. MFCs are arranged in series [37]. The reversal in some of the cells in the series stack is caused by the unavoidable increase in the internal resistances of the MFC units operated in series, as previously reported [37,38]. Thereby, power performance is reduced. When operated in parallel however, if the impedances of the MFCs are well matched, then the internal resistance of the MFC stack will tend towards the lowest common denominator and thus be more uniform [39]. This is evident by the reduction in internal resistance of the MFC_S stack from 244 to 76 kΩ. This large reduction in the internal resistance may also explain the increase in the current density of the parallel stack from 7.3 mA m -2 to 18.4 mA m -2 , as summarised in Table 1.
Considering the results obtained for the MFC_S stacks, the MFC_L devices were arranged only in parallel. As shown in Figure 5B, in this case the maximum power output of the stack was nearly 6 times higher compared to the MFC_L individual units. The power density increased by a factor of 2, and the internal resistance decreased from 33 kΩ to 1.4 kΩ ( Table   1).
The stacking of larger MFCs (mL scale) has been shown to increase the power density of MFCs, albeit not to the extent observed in this report. For example power densities of millilitre scale MFCs (6.25 and 12 mL) were improved by a factor of 1.2-1.4 by stacking multiple units together [9,38]. On the other hand,  demonstrates similar power densities between individual units and MFC stacks when using 60 mL MFCs.

Use of Biomass-Derived ORR Catalysts
To enhance power generation, without compromising cost-effectiveness and sustainability, two biomass-derived carbon materials, BC1 and BC2, were tested as ORR catalysts at the cathode. Since MFC_L showed better performance, this study was carried out only on this fuel cell design. The resulting fuel cells were named as MFC_BC1 and MFC_BC2 according to the type of catalyst used. Table 1 summarises the results obtained and compares them with the catalyst-free fuel cells previously tested. Figure 6 shows the polarisation and power curves for both devices. The OCV values for MFC_BC1 and MFC_BC2 were 151 mV and 220 mV respectively, and thus comparable with MFC_L.
As expected, the ORR catalysts enhanced the power performance of the MFCs, leading to a power output and power density almost 3 times higher than MFC_L. The effectiveness of biomass-derived ORR catalysts may be attributed to the large surface area [19] that the materials exhibit on the cathode surface compared to the plain carbon cloth (BC1: 376 m 2 g -1 ), as well as the capacity of heteroatom doping, such as nitrogen and sulphur, or the incorporation of nanoparticles like iron within the catalyst material to enhance the ORR activity [17,18,[41][42][43][44].
The internal resistances decreased to values of 15 kΩ and 23 kΩ, for MFC_BC1 and MFC_BC2 respectively, down to half those of MFC_L. Consequently, the current densities were an order of magnitude higher, with a value as high as 127.6 mA m -2 for MFC_BC1.
Generally MFC_BC1 performed better, with a 13% higher power density and a 44% increase in current density, compared to MFC_BC2. The structure of the two doped cathodes may be the reason for this difference. From the SEM images of the doped cathodes (Figure 7), it can be seen that the two ORR catalysts led to very different surface structures. In particular, it appears that BC1 percolated between the carbon fibres of the carbon cloth. Hence, good contact was formed between the carbon fibre electrode and the biomass-derived ORR catalyst, thus allowing a good active surface area for oxygen reduction reactions at the cathode surface.
On the other hand, BC2 formed a porous layer on top of the carbon fibres, which have resulted in an added resistance to the system and may explain the poorer performance of MFC_BC2 with respect to MFC_BC1.

Conclusions
Microbial fuel cells are an extremely attractive technology for the generation of clean electricity from a range of waste streams. The most viable route to boosting power density generated by MFCs is to develop small scale devices and arrange multiple units in stacks.
In this context, our study aims to guide towards the development of effective miniature MFCs. For this purpose we have developed an innovative miniature MFC, which can easily be further miniaturised. We have used an air-cathode configuration since it has the advantage of greater operational simplicity and cost-effectiveness. While fixing the electrodes spacing to 4 mm, we have investigated the effect of the electrodes length, when the system was continuously fed with artificial urine at a fixed flow rate of 0.36 mL min -1 .
The doubling of the electrode length of the miniature MFC, and so the hydraulic retention time as well, increased the power density more than tenfold due to enhanced mass transfer properties and substrate consumption at the electrode surface.