**3. Result and discussion**

Microorganism can be used in MFCs to catalyze the conversion of organic matter into electricity. The performance of the MFC was evaluated by the polarization curve and power density. The main goal of research to work on MFC is to increase output power and receive maximum generated current under optimum potential conditions.

Polarization behavior of the fabricated cell was recorded for several external resistances to determine maximum power generation. Polarization curve and power density vs. current density of the cell after 12 hours incubation and also reaching to steady state (SS) condition are presented in Fig. 3. The maximum produced power without any electron shuttle in anode was 4 mW.m-2. The produced power and current were very low to use in a small device and it must be improved.

Mediators are normally used to enhance the performance of MFCs (Najafpour et al.). Mediators are artificial compounds or produced by the microorganism itself. Some microorganisms produce nanowires to transmit electrons directly without using any mediator but other organisms need to add artificial electron shuttle into anode chamber (Mathuriya and Sharma, 2009). Yeast cannot transfer the produced electrons to the anode surface without addition of mediators. In orther to improve the power density and also current density several mediators with several concentrations were selected to enhance the power generation and current in the fabricated MFC. The maximum power, maximum current and also the obtained OCV at the best concentration of each mediator are summarized in Table 2. The data indicated that the mediators were essential when yeast was used as active biocatalyst in the MFC. Also this table indicated NR with concentration of 200 µmol.l-1 had the best ability for transferring the generated electrons in the anode chamber to the anode surface. The indicated concentration of NR in anaerobic anode compartment increased the produced power was 46 times more than the case without mediators in the MFC.


Table 2. Optimum condition obtained from this study at several concentrations of mediators

electrode were joined together to measure oxidation and reduction peaks. Carbon paper (NARA, Guro-GU, Seoul, Korea) was used as the working electrode and Platinum (Platinum, gauze, 100 mesh, 99.9% meta basis, Sigma Aldrich) as the counter electrode. Also, Ag/AgCl (Ag/AgCl, sat KCl, Sensortechnik Meinsberg, Germany) electrode was utilized as

Microorganism can be used in MFCs to catalyze the conversion of organic matter into electricity. The performance of the MFC was evaluated by the polarization curve and power density. The main goal of research to work on MFC is to increase output power and receive

Polarization behavior of the fabricated cell was recorded for several external resistances to determine maximum power generation. Polarization curve and power density vs. current density of the cell after 12 hours incubation and also reaching to steady state (SS) condition are presented in Fig. 3. The maximum produced power without any electron shuttle in anode was 4 mW.m-2. The produced power and current were very low to use in a small

Mediators are normally used to enhance the performance of MFCs (Najafpour et al.). Mediators are artificial compounds or produced by the microorganism itself. Some microorganisms produce nanowires to transmit electrons directly without using any mediator but other organisms need to add artificial electron shuttle into anode chamber (Mathuriya and Sharma, 2009). Yeast cannot transfer the produced electrons to the anode surface without addition of mediators. In orther to improve the power density and also current density several mediators with several concentrations were selected to enhance the power generation and current in the fabricated MFC. The maximum power, maximum current and also the obtained OCV at the best concentration of each mediator are summarized in Table 2. The data indicated that the mediators were essential when yeast was used as active biocatalyst in the MFC. Also this table indicated NR with concentration of 200 µmol.l-1 had the best ability for transferring the generated electrons in the anode chamber to the anode surface. The indicated concentration of NR in anaerobic anode compartment increased the produced power was 46

> Pmax (mW.m-2)

mediators --- 0.8 11 280

Ferric chelate 400 7.3 67 285

Thionine 500 12 79 460

NR 200 37 151 505

MB 300 8.3 71 410

Table 2. Optimum condition obtained from this study at several concentrations of mediators

I max in Pmax (mA.m-2)

OCV at SS condition (mV)

reference electrode. Voltage rate of 50 mV.S-1 was chosen as scan rate in CV analysis.

maximum generated current under optimum potential conditions.

times more than the case without mediators in the MFC.

Optimum concentration (µ mol.l-1)

**3. Result and discussion** 

device and it must be improved.

Type of mediators

Without

Fig. 3. Generated power density (a) and voltage (b) as function of current density at start up, 10 hours after incubation and at steady state condition

Effect of Mass Transfer on Performance of Microbial Fuel Cell 241

Nafion area: 3.14 cm2 Nafion area: 9cm2 Nafion area: 16 cm2

Voltage (mV)

Power (mW.m-2)

0

20

40

60

80

100

120

140

160

0

200

400

600

800

1000

Current (mA.m-2) 0 200 400 600 800 1000

Current (mA.m-2)

(b)

0 200 400 600 800

Nafion area: 3.14 cm2 Nafion area: 9 cm2 Nafion area: 16 cm2

Fig. 5. Effect of mass transfer area on performance of MFC.

(a)

In order to obtain the best oxidizer in cathode compartment, several oxidizers were analyzed. Table 3 summarized the optimum conditions obtained for distilled water, potassium ferricyanide and potassium permanganate. The maximum power, current and OCV was obtained with potassium permanganate.


Table 3. Optimum conditions obtained from several oxidizers

Glucose consumption and cell growth with respect to incubation time at 200µmol.l-1 of NR as electron mediators are presented in Fig. 4. Figure 4 demonstrated that *S. cerevisiae* had the good possibility for consumption of organic substrate at anaerobic condition and produce bioelectricity.

The aim of this research was to found optimum effect of mass transfer area on production of power in the fabricated MFC. Figure 5 shows the effect of mass transfer area on performance

Fig. 4. Cell growth profiles and glucose consumption by *S. cerevisiae*

In order to obtain the best oxidizer in cathode compartment, several oxidizers were analyzed. Table 3 summarized the optimum conditions obtained for distilled water, potassium ferricyanide and potassium permanganate. The maximum power, current and

Distillated water --- 7.6 68 404

H2O2 --- 41 155 610

ferricyanide 200 49 177 508

Permanganate 300 110 380 860

Glucose consumption and cell growth with respect to incubation time at 200µmol.l-1 of NR as electron mediators are presented in Fig. 4. Figure 4 demonstrated that *S. cerevisiae* had the good possibility for consumption of organic substrate at anaerobic condition and produce

The aim of this research was to found optimum effect of mass transfer area on production of power in the fabricated MFC. Figure 5 shows the effect of mass transfer area on performance

> Time (h) 0 10 20 30 40 50 60

Glucose consumption

OD

Fig. 4. Cell growth profiles and glucose consumption by *S. cerevisiae*

Pmax (mW.m-2)

I max in Pmax (mA.m-2) OCV at SS condition (mV)

Absorbance at 620 nm

0.0

0.2

0.4

0.6

0.8

1.0

1.2

OCV was obtained with potassium permanganate.

Optimum concentration (µ mol.l-1)

Table 3. Optimum conditions obtained from several oxidizers

Type of Oxidizer

Potassium

Potassium

bioelectricity.

Glucose concentration (g.l-1)

0

5

10

15

20

25

30

35

Fig. 5. Effect of mass transfer area on performance of MFC.

Effect of Mass Transfer on Performance of Microbial Fuel Cell 243

was continuously injected from feed tank to the anode chamber using a peristaltic pump. Four different HRT were examined in this research to determine the optimum HRT for maximum power and current density. The polarization curve at each HRT at steady state condition was recorded with online data acquisition system and the obtained data are presented in table 4. The optimum HRT was 6.7 h with the maximum generated power

16 161 420 801 12.34 182 600 803 6.66 274 850 960 3.64 203 614 975 Table 4. Effect of different HRT on production of power and current in fabricated MFC

The growth kinetics and kinetic constants were determined for continuous operation of the fabricated MFC. The growth rate was controlled and the biomass concentration was kept constant in continuous system through replacing the old culture by fresh media. The material balance for cells in a continuous culture is defined by equation 5 (Bailey and Ollis,

�� �� � �� � � �� �� = �� ��

where, F is volumetric flow rate of feed and effluent liquid streams, V is volume of liquid in system, rx is the rate of cell growth, xi represents the component i molar concentration in feed stream and x is the component i molar concentration in the reaction mixture and in the effluent stream. The rate of formation of a product is easily evaluated at steady-state condition for inlet and outlet concentrations. The dilution rate, D, is defined as D=V/F which characterizes the inverse retention time. The dilution rate is equal to the number of fermentation vessel volumes that pass through the vessel per unit time. D is the reciprocal of

At steady-state condition, there is no accumulation. Therefore, the material balance is

When feed is steriled, there is no cell entering the bioreactor, which means x0=0. Using the Monod equation for the specific growth rate in equation 6, the rate may be simplified and

�� � = μ� = (��� �� �)

HRT (h) 16 12.34 6.66 3.64 X (g.l-1) 1.94 1.74 1.728 1.5 S(g.l-1) 6.95 9.13 12.86 22.8 Table 5**.** Biomass and substrate concentration in outlet of MFC at different HRT

�� = (� � ��)

�� �� � �� � � �� �� = 0 → �

�� = ��

I max in Pmax (mA.m-2)

OCV at SS condition (mV)

�� � (5)

�� � (6)

�� � � � (7)

Pmax (mW.m-2)

density of 274 mW.m-2.

1976):

reduced to:

HRT (h)

the mean residence time(Najafpour, 2007).

reduced to following equation:

of MFC. Three different mass transfer area (3.14, 9and 16 cm2) were experimented and the results in polarization curve presented in Fig. 5 a and b. Membrane in MFC allows the generated hydrogen ions in the anode chamber pass through the membrane and then to be transferred to cathode chamber (Rabaey et al., 2005a; Cheng et al., 2006; Venkata Mohan et al., 2007; Aelterman et al., 2008). The obtained result shows the maximum current and power were obtained at Nafion area of 16 cm2. The maximum power and current generated were 152 mW.m-2 and 772 mA.m-2, respectively.

Figure 6 depicts an OCV recorded by online data acquisition system connected to the MFC for duration of 72 hours. At the starting point for the experimental run, the voltage was less than 250mV and then the voltage gradually increased. After 28 hours of operation, the OCV reached to a maximum and stable value of 8mV. The OCV was quite stable for the entire operation, duration of 72 hours.

Fig. 6. Stability of OCV.OCV recorded by online data acquisition system connected to the MFC for duration of 72 hours

There are several disadvantages of batch operation for the purpose of power generation in MFCs. The nutrients available in the working volume become depleted in batch mode. The substrate depletion in batch MFCs results in a decrease in bioelectricity production with respect to time. This problem is solved in continuous MFCs that are more suitable than batch systems for practical applications (Rabaey et al., 2005c). The advantages of continuous culture are that the cell density, substrate and product concentrations remain constant while the culture is diluted with fresh media. The fresh media is sterilized or filtered and there are no cells in the inlet stream.

The batch operation was switched over to continuous operation mode by constantly injection of the prepared substrate to the anode compartment. The other factors were kept constant based on optimum conditions determined from the batch operation. For the MFC operated under continuous condition, substrate with initial glucose concentration of 30 g.l-1

of MFC. Three different mass transfer area (3.14, 9and 16 cm2) were experimented and the results in polarization curve presented in Fig. 5 a and b. Membrane in MFC allows the generated hydrogen ions in the anode chamber pass through the membrane and then to be transferred to cathode chamber (Rabaey et al., 2005a; Cheng et al., 2006; Venkata Mohan et al., 2007; Aelterman et al., 2008). The obtained result shows the maximum current and power were obtained at Nafion area of 16 cm2. The maximum power and current generated

Figure 6 depicts an OCV recorded by online data acquisition system connected to the MFC for duration of 72 hours. At the starting point for the experimental run, the voltage was less than 250mV and then the voltage gradually increased. After 28 hours of operation, the OCV reached to a maximum and stable value of 8mV. The OCV was quite stable for the entire

> Time (h) 0 20 40 60 80

Fig. 6. Stability of OCV.OCV recorded by online data acquisition system connected to the

There are several disadvantages of batch operation for the purpose of power generation in MFCs. The nutrients available in the working volume become depleted in batch mode. The substrate depletion in batch MFCs results in a decrease in bioelectricity production with respect to time. This problem is solved in continuous MFCs that are more suitable than batch systems for practical applications (Rabaey et al., 2005c). The advantages of continuous culture are that the cell density, substrate and product concentrations remain constant while the culture is diluted with fresh media. The fresh media is sterilized or filtered and there are

The batch operation was switched over to continuous operation mode by constantly injection of the prepared substrate to the anode compartment. The other factors were kept constant based on optimum conditions determined from the batch operation. For the MFC operated under continuous condition, substrate with initial glucose concentration of 30 g.l-1

OCV

were 152 mW.m-2 and 772 mA.m-2, respectively.

operation, duration of 72 hours.

Voltage (mV)

200

MFC for duration of 72 hours

no cells in the inlet stream.

300

400

500

600

700

800

900

1000

was continuously injected from feed tank to the anode chamber using a peristaltic pump. Four different HRT were examined in this research to determine the optimum HRT for maximum power and current density. The polarization curve at each HRT at steady state condition was recorded with online data acquisition system and the obtained data are presented in table 4. The optimum HRT was 6.7 h with the maximum generated power density of 274 mW.m-2.


Table 4. Effect of different HRT on production of power and current in fabricated MFC

The growth kinetics and kinetic constants were determined for continuous operation of the fabricated MFC. The growth rate was controlled and the biomass concentration was kept constant in continuous system through replacing the old culture by fresh media. The material balance for cells in a continuous culture is defined by equation 5 (Bailey and Ollis, 1976):

$$\left.F.\varkappa\_l - F.\varkappa + V.\tau\_\mathbf{x} = V.\begin{pmatrix}d\mathbf{x} \\ dt\end{pmatrix}\right|\_{\mathbf{d}\mathbf{t}}\tag{5}$$

where, F is volumetric flow rate of feed and effluent liquid streams, V is volume of liquid in system, rx is the rate of cell growth, xi represents the component i molar concentration in feed stream and x is the component i molar concentration in the reaction mixture and in the effluent stream. The rate of formation of a product is easily evaluated at steady-state condition for inlet and outlet concentrations. The dilution rate, D, is defined as D=V/F which characterizes the inverse retention time. The dilution rate is equal to the number of fermentation vessel volumes that pass through the vessel per unit time. D is the reciprocal of the mean residence time(Najafpour, 2007).

At steady-state condition, there is no accumulation. Therefore, the material balance is reduced to:

$$\left\langle F.\,\boldsymbol{x}\_{l}-F.\,\boldsymbol{x}+V.\,\boldsymbol{r}\_{\boldsymbol{x}}=\boldsymbol{0} \quad\rightarrow \,^{V}\rangle\_{F} = \left\langle \boldsymbol{\pi}-\boldsymbol{x}\_{0} \right\rangle\_{\mathcal{T}\_{\mathbf{x}}} \tag{6}$$

When feed is steriled, there is no cell entering the bioreactor, which means x0=0. Using the Monod equation for the specific growth rate in equation 6, the rate may be simplified and reduced to following equation:

$$
\pi\_{\mathfrak{x}} = \left. ^{d\mathfrak{x}}\right\rangle\_{\mathfrak{tt}} = \mathfrak{\mu}\mathfrak{x} = \left. ^{\{\mu\_{\max}, \mathcal{S}, \mathfrak{x}\}}\right\rangle\_{\mathcal{K}\_{\mathfrak{s}} + \mathcal{S}} \tag{7}
$$


Table 5**.** Biomass and substrate concentration in outlet of MFC at different HRT

Effect of Mass Transfer on Performance of Microbial Fuel Cell 245

Biomass and substrate concentration in outlet stream of MFC at different HRT are shown in Table 4. To evaluate kinetic parameters, the double reciprocal method was used for linearization. The terms µmax and Ks were recovered from a linear fit of the experimental data by Plotting 1/D versus 1/S. The values obtained for µmax and Ks were 0.715 h and 59.74

In the next stage, anode electrode with attached microorganisms was analyzed with CV in. The system was analyzed in anaerobic anode chamber. Before formation of active biofilm on anode surface, oxidation and reduction peak was not observed in CV test (Fig. 7a). Currentpotential curves by scanning the potential from negative to positive potential after formation of active biofilm are shown in Fig. 7b. Two oxidation and one reduction peak was obtained with CV test. One peak was obtained in forward scan from -400 to 1000 mV and one oxidation and reduction peak was obtained in reverse scan rate from 1000 to -400 mV. The similar result by alcohol as electron donors in anode chamber was reported(Kim et al., 2007). The first peak was observed in forward scan rate between -0.087 to 1.6 V. Also 200 mol.l-1 NR was added to

Graphite was used as electrode in the MFC fabricated cells. The normal photographic image of the used electrode before employing in the MFC as anode compartment is shown in Fig. 8a. Scanning electronic microscopy technique has been applied to provide surface criteria and morphological information of the anode surface. The surface images of the graphite plate electrode were successfully obtained by SEM. The image from the surface of graphite electrode before and after experimental run was taken. The sample specimen size was 1cm×1cm for SEM analysis. Fig. 8b and 8c show the outer surface of the graphite electrode prior and after use in the MFC, respectively. These obtained images demonstrated that microorganisms were grown on the graphite surface as attached biofilm. Some clusters of

����� � � � (8)

�� <sup>=</sup> ��������� ��

g/l, respectively. Then, the kinetic model is defined as follows:

anode chamber and then this system was examined with CV (Fig. 7 c)

microorganism growth were observed in several places on the anode surface.

(a) (b) (c)

after (c) using in anode compartment

summarized below:

Fig. 8. Photography image (a) and SEM images from anode electrode surface before (b) and

Yeast as biocatalyst in the MFC consumed glucose as carbon source in the anode chamber and the produced electrons and protons. In this research, glucose was used as fuel for the MFC. The anodic and catholic reactions are taken place at the anode and cathode as

Fig. 7. Effect of active biofilm on anode surface with CV analysis. (a) absence of biofilm ,(b) after formation of biofilm with out mediators and (c) after formation of biofilm with 200 µmol.l-1 NR as electron mediators .scan rate was 0.01 V.S-1

Current (mA) -0.8 -0.6 -0.4 -0.2 0.0 0.2 0.4 0.6 0.8 1.0

Current (mA)


Current (mA)


µmol.l-1 NR as electron mediators .scan rate was 0.01 V.S-1


0.0

0.5

1.0

1.5


0.0

0.5

1.0

1.5

Potential (V) -0.6 -0.4 -0.2 0.0 0.2 0.4 0.6 0.8 1.0 1.2

(a)

Potential (V) -0.6 -0.4 -0.2 0.0 0.2 0.4 0.6 0.8 1.0 1.2

(b)

pottential (V) -0.6 -0.4 -0.2 0.0 0.2 0.4 0.6 0.8 1.0 1.2

(c) Fig. 7. Effect of active biofilm on anode surface with CV analysis. (a) absence of biofilm ,(b) after formation of biofilm with out mediators and (c) after formation of biofilm with 200

Biomass and substrate concentration in outlet stream of MFC at different HRT are shown in Table 4. To evaluate kinetic parameters, the double reciprocal method was used for linearization. The terms µmax and Ks were recovered from a linear fit of the experimental data by Plotting 1/D versus 1/S. The values obtained for µmax and Ks were 0.715 h and 59.74 g/l, respectively. Then, the kinetic model is defined as follows:

$$r\_{\chi} = \binom{0.715 \text{ S.x}}{59.74 + \text{ S}} \tag{8}$$

In the next stage, anode electrode with attached microorganisms was analyzed with CV in. The system was analyzed in anaerobic anode chamber. Before formation of active biofilm on anode surface, oxidation and reduction peak was not observed in CV test (Fig. 7a). Currentpotential curves by scanning the potential from negative to positive potential after formation of active biofilm are shown in Fig. 7b. Two oxidation and one reduction peak was obtained with CV test. One peak was obtained in forward scan from -400 to 1000 mV and one oxidation and reduction peak was obtained in reverse scan rate from 1000 to -400 mV. The similar result by alcohol as electron donors in anode chamber was reported(Kim et al., 2007). The first peak was observed in forward scan rate between -0.087 to 1.6 V. Also 200 mol.l-1 NR was added to anode chamber and then this system was examined with CV (Fig. 7 c)

Graphite was used as electrode in the MFC fabricated cells. The normal photographic image of the used electrode before employing in the MFC as anode compartment is shown in Fig. 8a. Scanning electronic microscopy technique has been applied to provide surface criteria and morphological information of the anode surface. The surface images of the graphite plate electrode were successfully obtained by SEM. The image from the surface of graphite electrode before and after experimental run was taken. The sample specimen size was 1cm×1cm for SEM analysis. Fig. 8b and 8c show the outer surface of the graphite electrode prior and after use in the MFC, respectively. These obtained images demonstrated that microorganisms were grown on the graphite surface as attached biofilm. Some clusters of microorganism growth were observed in several places on the anode surface.

Fig. 8. Photography image (a) and SEM images from anode electrode surface before (b) and after (c) using in anode compartment

Yeast as biocatalyst in the MFC consumed glucose as carbon source in the anode chamber and the produced electrons and protons. In this research, glucose was used as fuel for the MFC. The anodic and catholic reactions are taken place at the anode and cathode as summarized below:

Effect of Mass Transfer on Performance of Microbial Fuel Cell 247

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$$\text{C}\_6\text{H}\_2\text{O}\_6 + 6\text{H}\_2\text{O} \xrightarrow[-]{} \text{D} + 6\text{CO}\_2 + 24\text{ e} + 24\text{H}^+\tag{9}$$

$$2\,\text{6O} \,\text{2} + 2\,\text{4} \,\text{e} + 2\,\text{4H} \,\text{4} \xrightarrow[]{} \text{12H} \,\text{5O} \tag{10}$$

24 mol electrons and protons are generated by oxidation of one mole of glucose in an anaerobic condition. To determine CE (Columbic Efficiency), 1 KΩ resistance was set at external circuit for 25 h and the produced current was measured. The average obtained current was 105.85 mA.m-2. In this study, CE was calculated using equations 3 and 4. CE was 26% at optimum concentration of NR as mediator. CE at continues mode was around 13 percent and this efficiency is considered as very low efficiency. The similar results with xylose in fed-batch and continuous operations were also reported (Huang and Logan, 2008b; a). This may be due to the breakdown of sugars by microorganisms resulting in production of some intermediate products such as acetate, butyrate, and propionate, which can play a significant role in decrease of CE.
