BIO301 Chemostat Group Report

Generation of Electricity by Mediatorless Microbial Fuel Cell using Bacteria Source from Activated Sludge

Lester Fong, Tony Ho, Ankit Broker, David Hii, Yousif Hanna, Jeremy Hartley &

James Atem.

Abstract

Microbial fuel cells generate electricity by harnessing the electron transport chain of

bacteria under controlled conditions., Tthe basic design of most microbial fuel cells

consists of an anaerobic anode chamber containing feed source and inoculated with a

mixed microbial culture and an anode cathode chamber which contains an oxidizing

agent such as dissolved oxygen of ferricyanide. The power output of a microbial fuel

cell was measured in terms of a polarisation curve, which shows the relationship

between current and voltage over a range of resistances. The polarisation curves were

performed for with two different methods. Firstly, when the MFC were in batch

culture the maximum voltage obtained for the first set of results was 0.272V and the

second set of results revealed 0.278V indicating bacteria had increased in health. The

couloumbic efficiency in the batch was recorded at 0.665%, which revealed the

bacteria were extremely weak. The first power curve recorded was at 0.0035 mW at a

current of 0.0372 mA which was rather low. Furthermore the second power curve

showed a maximal power output of 0.0089 mW at a current of 0.063 mA revealing the

bacteria were healthier as the maximal power was higher and at a lower resistance

compared to the previous one. Resistance kept decreasing when the power was at its

maximum. Finally when the MFC were was operated introduced in as a continuous

culture (chemostat), the maximum voltage obtained from both sets of chemostat

curves was 0.556V, which showed an increase in the voltage when compared to the

batch culture (0.278V & 0.272V). It is a bit inconsistent first evaluating the fuel cell

by power output (mW) and then switch to volts. If volts are used the resistance needs

to be specified otherwise it does not mean much.

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Introduction

Microbial fuel cells (MFC’s) generate electricity by harnessing the electron transport

chain of bacteria under controlled conditions (Mohan et al., 2007). They have

potential to generate electricity from a wide variety of organic wastes while oxidising

the wastes to less harmful forms (Moon et al., 2006, Ieropoulos et al., 2005, Liu et al.,

2004). Research into developing efficient MFC’s remains a very current field

(unclear) , with both engineering and biological challenges yet to be met (Moon et al.,

2006, Mohan et al., 2007).

The basic design of most microbial fuel cells consists of an anaerobic anode chamber

containing a feed source and inoculated with a mixed microbial culture, and an anode

cathode chamber which contains an oxidizing agent such as dissolved oxygen of

ferricyanide (Mohan et al., 2007). The use of cut and paste for sentences is not a

good idea, in particular if the same mistake (refer to abstract) is made (confusing

anode with cathode). Being deprived of a direct electron acceptor for respiration, the

bacteria in the anode chamber donate electrons to the anode, which are then

transferred via a conductor to the cathode, where reduction (of what?) occurs

(Ieropoulos et al., 2005). Charge balance is maintained by migration of H+ across a

proton exchange membrane (Mohan et al., 2007).

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Figure 1: Schematic representation of a mediatorless (it has still not been

explained what this means) MFC operating in batch mode using acetate as an

electron donor and ferricyanide as an electron acceptor.

MFC’s may be broadly classified into two categories depending on the means of

electron transfer between the bacteria and the anode. Mediated fuel cells contain an

artificial mediator in the anode chamber (Liu et al., 2004). (good, to have it explained

now) Bacteria transfer electrons to the mediator in solution, which is then regenerated

at the anode (Ieropoulos et al., 2005, Liu et al., 2004). This mechanism of electron

transfer has several disadvantages relating to the cost and toxicity of artificial

mediators (Ieropoulos et al., 2005, Mohan et al., 2007, Liu et al., 2004). A second

category of MFC’s does not contain an artificial mediator, but relies on natural

electron transfer processes of the bacteria. While these processes are as yet poorly

understood, they are thought to include direct electron transfer by membrane bound

enzymes as well as synthesesis of natural mediators (Ieropoulos et al., 2005, Stams et

al., 2006, Liu et al., 2004).

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Because not all substrate is completely oxidised, with some mass necessarily being

used for biosynthesis, then not all high energy electrons supplied in the substrate are

transferred to the cathode and available to do work. The percentage of electrons which

are transferred is expressed in terms of couloumbic efficiency, which is essentially a

percentage ratio of the number of electrons supplied as the organic substrate against

the number of electrons transferred. This parameter is a useful measure of the overall

efficiency of the MFC (Liu et al., 2004, Min and Logan, 2004, Williams, 1966).

(Good)

The power output of a MFC is also a useful quantity to measure. This is measured in

terms of a polarisation curve, which shows the relationship between current and

voltage over a range of resistances(Mohan et al., 2007, Larminie and Dicks, 2003). By

the relationships V=IR and P=IV, where V is voltage, I is current, R is resistance, and

P is power, then observations of current, voltage and resistance can be manipulated to

give information about power output (Atkins and de Paula, 2006, Serway and Faughn,

2003).

The aim of this experiment shall be to create a MFC which is capable of degrading

wastewater to produce electricity, and to investigate it’s performance under both batch

and continuous flow conditions.

(The intro is pretty good, finishing by introducing the aim. Could attempt to refer to

the fact that this is supposed to be a chemostat project).

Materials and Methods

Microbial Fuel cell setup

The fuel cell consisted of two chambers (each 500 mL), an anode chamber and a cathode chamber, which were separated by a proton exchange membrane between solutions, and a conductor between the electrodes. The anode consisted of a carbon sponge connected to a platinum wire, while the cathode was composed of platinum

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foil.(Figure 9).The anode chamber was inoculated with activated sludge and kept under anaerobic conditions, while the cathode chamber was filled with 500mL Potassium Ferricyanide and kept under aerobic conditions. (Figure 10).Both chambers were stirred magnetically and maintained in a water bath at 30oC. The circuit between electrodes was closed to allow electron transfer. (Figure 2). The solution in the cathode chamber was changed when the fading colour of the solution indicated near complete reduction of the Ferricyanide.

(It would be good to see a schematic drawing)

Operation

Batch mode:

Synthetic wastewater was prepared with acetate as a carbon source and electron donor. At this stage of the experiment the acetate was kept as a separate solution to limit microbial contamination. The acetate solution was prepared to a concentration of 1.00 M , and the composition of the other component of the synthetic wastewater is given below):

1L Synthetic wastewater consists 20mL of sodium acetate 1.0M, 1.25mL trace elements, 480mg NaHCO3, 95.5mg NH4Cl, 10.5 K2HPO4, 5.25 KH2PO4, 63.1 CaCl2.2H20, and 19.2 MgSO4.7H20 (Ghangrekar and Shinde, 2006).

Chemostat mode:

Synthetic wastewater was supplied to the anode chamber at a rate of 50 mL per day via a peristaltic pump running on a timer cycle of 288 minutes off, 1 minute on. (It is strange to run a chemostat essentially as a sequencing batch system, 288 min off is almost 5 hours. It would be better to use seconds instead of minutes) The substrate reservoir contained a single part AWW including diluted acetate component. It was kept in an insulated icebox to reduce microbial activity, and stirred magnetically and intermittently on a timer cycle of 0.5 minutes every 70 minutes to increase homogeneity but minimise aeration. (Figure 11). (Explanations are nice)

Monitoring of voltage

A Voltmeter was used to measure the potential against 1000ohm and the values were recorded down periodically.

Polarization curves

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Polarisation curves were used as a means of determining the capacity of the bacterial culture. (Would be nice to have some further explanations why polaristion curves are needed.) A substrate saturated culture was used, and voltage was measured at a series of stepwise increasing resistances until no voltage was measurable between the electrodes. Using the relationship V=IR, then current was thus calculated at each point of the curve, allowing power to also be calculated by the relationship P=IV (OK, but…. Show units to allow next years students an easier start)

Methods for obtaining polarization curve

The resistor was disconnected and the potential was allowed to build up to a point where no further increment of voltage can be observed. This was followed by observation of voltage increment using voltmeter until it reaches the maximum potential point. Then, the resistor was connected and ensured that the connection is a close circuit system as illustrated below:

Figure 2: Closed circuit system of microbial fuel cell

The potential values were recorded while varying the resistance from the highest to the lowest resistance at time intervals of 5 minutes. (Note: The voltage values must be taken only when the pseudo-steady-state (explain what it is and why it is necessary)

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conditions have been established).Graphs of cell cottage (V) versus current (mA) and Power (mW) versus current (mA) were plotted by referring to the recorder values.

Methods for obtaining Columbic (it was not the Spanish conquerer Columbus but the French Coulomb) efficiency in batch culture

The resistor was connected to between the anode and cathode chambers to establish a closed circuit system (As illustrated in figure 4 (Figure missing?) ). After that, the potential was measured using a voltmeter. Tthe microbial fuel cell was left to run overnight for the voltage reading of the potential to reach stabilization. Then, 1mL sodium acetate 1.0M was added to the anode chamber using Terumu needle (0.65 x 32mm) with syringe. After addition, the voltage reading from the voltage meter was recorded at an interval of 2 hours (Take note (You need to refer to the figure that shows this) of the highest possible voltage reading before the curve start to drop – refer results) and eventually stabilize.

Stirring speed

When Speed

During batch culture 220rpm

1st polarization curve (batch mode) 360rpm

2nd polarization curve (batch mode) 360rpm

3rd polarization curve (chemostat) 200rpm

4rd polarization curve (chemostat) 200rpm

Temperature

The temperature of the water in aquarium was maintained at around 30°C (this is to ensure suitable temperature for the bacteria to grow in the anode chamber). The ice in the Esky Box was changed 2 times per day (this prevents contamination due to growth of other microorganisms)

pH

pH of the anode chamber was maintained at around pH6.5 – pH7.0.

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Results

Each result is being first introduced, then explained and interpreted. This needs at

least one paragraph. Also before a figure is shown, the text needs to refer to it (as in

the journal papers that you have been reading) Initially, results were taken when the

MFC was in a batch culture.

Figure 3: Batch polarization curves.

From such calculations, a variety of curves were produced. The polarisation curves

produced showed the effects of voltage on the current (Fig 3). Such a graph shows the

lowering of the potential of an electrode from equilibrium, which is caused by the

passing of an electric current. On the other hand, a power curve illustrates the highest

amount of power that the bacteria could produce, showing that the higher the

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maximum power, the ‘stronger’ the bacteria were. The other main reason for a power

curve is that, not only does it show that maximum potential output of the MFC but

also at what resistance. (You start talking about power curve. The reader would now

expect that you show the curve)t.

For the first polarisation curve, the higher the current, the greater the drop was in the

voltage (maximal velocity of 0.2724V) (Fig 3). Although it was not really visible

from the graph (refer to table 1), at the beginning (peak voltage), there was a slight

increase and then decrease in voltage. This area is where activation losses are

dominant (voltage area between 0.2725V and 0.2704V). Furthermore, ohmic losses

were dominant when the voltage was between 0.2704V and 0.2664V. When there was

a rapid, linear decrease in the voltage, at higher currents (between 0.2664V and about

0.0103V), concentration loss (mass transport effect) was overriding. (I am missing

some more simple observations such as: there was increased voltage or current after

acetate addition, indicating that acetate caused some electricity flow.)

The second polarisation curve was produced according to results that were taken three

days after the initial set of results (Fig 3). As seen in the previous results, the higher

the current, the greater the drop was in the voltage (Fig 3; Table 2). A higher maximal

voltage was achieved, which was 0.278V compared to 0.2724V. Activation losses did

not really dominate at all in this polarisation curve. (Unclear to me) If there was any

domination (??) , it would have been between a voltage of 0.278V and 0.270V.

Ohmic losses (explain what that is) were dominant when the voltage was between

0.270V and 0.2438V. When there was a linear decrease in voltage (linear as a

function of time or current or resitor?) , concentration loss (mass transport effect (??

What specifically is meant? Write more simply and more clearly) was dominant at a

voltage between 0.2438V and 0.0055V. To compare with the first polarisation curve,

the second one slightly shifted to the right, showing higher voltages at higher currents

(0.097mA (0.0145V) compared to the first curve, which had a current of 0.097mA

and a voltage of 0.0032V). (Overall there is too much writing on the two polarisation

curves without actually saying what they prove in terms of improvement of the MFC.)

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Figure 4: Batch power curves.

The first power curve showed that the power of the bacteria had a maximal output

potential of 0.0035mW, which was rather low (closed circuit only, as open circuit

showed no power) (Fig 4). This power was achieved at a current of 0.03728mA. At

maximal power, also, the resistance was 2500 ohms. The maximal volumetric power

equals power (W) divided by metres cubed (volume (L) of anode) (Table 1). This

occurs at the maximal output potential and in this part of the experiment, it equalled

1.39x10-5 W/m3, which was also a very low number. The reason for the drop in power,

after it maximises, is due to an increase in ohmic loss and electrode over potentials

(short circuit situation).

The maximal power output achieved for the second power curve was higher than the

previous curve (a slight shift to the right) (Fig 4). The second power curve showed a

maximal power output of 0.0089mW at a current of 0.063mA. At the maximal power,

the resistance was 2200 ohms, showing that the bacteria had become slightly more

‘healthier’, as the maximal power was higher and at a lower resistance (compared to

the previous power results). The maximal volumetric power was 3.54x10-5 W/m3,

(good to see you use volumetric power, now you just need to compare to literature

values) which was also higher than the previous result of 1.39x10-5 W/m3,

emphasising vast bacterial improvement.

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In batch mode, the columbic efficiency curve was plotted with current (mA) versus

time (h). It represents the total amount of time required for the bacteria in the MFC to

completely metabolise 1mL of 1M acetate. Calculated values for the graph can be

seen in Table 5 of the appendix.

Figure 5: Current as a function of time (determines coulombic efficiency) (It

would be more useful to jut give the final acetate concentration)

This part of the experiment was carried out three days after the second set of batch

results were achieved. The resistance used was the one that gave maximal output

potential in the latest set of results (2200 ohms) (Fig 4).

It is known that, current = coulomb per time, while the area under the curve and above

the baseline = (Coulomb / Time) x Time = Coulomb. (Good to see this explanation, it

shows good understanding an is really useful) From Table 5, the area under the curve

and above the baseline was calculated to be 5.1 Coulombs using integration. Table 5

is a good example to show how the calculation works. It looks OK to me.

Calculating Columbic Efficiency %:

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Added 1mL Acetate (1M)

On addition of 1mL of 1M acetate

Number of moles of acetate added = 1/1000 x 1 = 0.001 moles of acetate

Number of moles of electrons available in 0.001 moles of acetate = 0.001 x 8 = 0.008 moles of e- (OK)

Since 1 moles of electrons = 96485 Coulombs, 0.008 moles of electrons = 96485 x 0.008 = 771.88 Coulombs. Hence, in theory, 0.001 moles of acetate should have 771.88 Coulombs. - (OK)

Columbic efficiency =

___________Coulombs calculated from the curve x 100__________

Theoretical amount of Coulombs in added amount of acetate

= 5.1 / 771.88 x 100 = 0.665% (which is an extremely low efficiency)

At a resistance of 2200 ohms, the voltage stabilised at 61.5mV (0.028mA), showing

that since the resistance is stable, an increase in voltage will cause an increase in

current (I=V/R) (Fig 5). Once the 1mL of acetate was added, a slow increase in

voltage occurred over about 7.2 hours, maxing out at 73mV (0.033mA). Once all the

acetate was metabolised, the voltage decreased over 13.8 hours, going back to the

baseline voltage of 61.5mV. The current baseline that was constructed over about 21

hours was used to find the area of the graph, which further assisted in calculating the

coulombic efficiency. (Describing the fig.)

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Figure 6: Correlation of acetate oxidation rate and current (Second batch

results).

The current shows the electron flow rate occurring in the MFC, which also equals the

current in coulombs (wow, you got the spelling right this time, It would have just

required a bit more thorough spell checking) per second (C/s). As known, one mole

of electrons (e-) equals 96485 C / mole e- (Faradays constant), therefore, to get the

current in moles of e-/sec it would be (C/s) divided by 96485. To get the moles of

acetate per hour it would be moles e- per second divided by 8 (acetate yields eight

electrons (requires two moles of oxygen)) x3600 (assuming all electrons are used to

make electricity). To get the acetate oxidation rate (moles of acetate/L/h), it would be

moles of acetate per hour divided by the volume of the anolyte (0.5L). This answer

should equal to the current (linear correlation), however, it was off by a factor of

about 111 in this experiment (Fig 6). The reason for this is due to the extremely low

coulombic efficiency (0.665%) (decrease coulombic efficiency, decreases oxidation

rate), as at low coulombic efficiencies, the chances of an accurate finding of the

acetate oxidation rate decreases severely. (the above chapter repeats to some extent

what was calculated earlier. Keep it concise )

Any speculations as to what has happened to the acetate?

The following day was when the MFC was when the culture was put in a continuous

culture (chemostat). (unclear sentence) 50mL of acetate (1M) was pumped into the

anode vessel every day (vessel total volume = 500mL). Further on, the dilution rate is

equal to the flow rate (L/h) divided by the volume (L). Therefore, the dilution rate

equals (0.05/24)/0.5, which equals 0.0042 h-1, and the average treatment time, also

known as the hydraulic retention time (HRT) equals 1/0.0042 h (240 h or 10 days).

(OK)

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Figure 7: Chemostat polarisation curves. (nice graphs (colours and fonts))

These results were taken five days after the chemostat was put into place (Fig 7; Table

3). As usual, an increase in current caused an eventual decrease in voltage. In the first

chemostat polarization curve, there was quite a high increase in the voltage compared

to the batch culture (0.453V compared to 0.278V and 0.2724V) (Fig 7; Fig 1).

Activation losses dominated at a voltage between 0.453V and 0.452V (may not have

even occurred), while ohmic losses surged at a voltage between 0.452V and 0.412V.

The linear drop in voltage (concentration loss) was evident when the voltage was

between 0.412V and 0.0335V. Once again, if this polarisation curve was added to the

other two, there would be a larger shift to the right and the graph would be starting at

a higher point, showing that the voltage stayed higher at a higher current for a longer

time, in comparison to the others.

The second set of chemostat results were obtained the following day. (The

experimental plan is not obvious A voltage of 0.556V occurred in the final set of

results and this was the healthiest stage of the bacteria in this experiment (previous

voltage of 0.453V) (Fig 7). Activation loss was not really evident but ohmic losses

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occurred at voltages between 0.557V and 0.535V, while concentration loss (??)

dominated at a voltage between 0.535V and 0.215V. Overall, it was clear to see that

the highest voltages, over various currents, occurred in the last set of results that were

conducted in this experiment.

Figure 8: Chemostat power curves.

A new maximal output potential was achieved, being about 13 times higher than the

previous maximum one (0.115mW compared to 0.0089mW) (Fig 8; Fig 4). This

occurred at a current of 0.45mA and at a resistance of 560 ohms, which was almost

2000 less ohms than the final batch culture results. This, once again, showed that the

bacteria were a lot stronger compared to the previous tests. The maximal volumetric

power was 3.54x10-5 W/m3, which was also higher than the previous maximum result

of 1.39x10-5 W/m3.

The highest maximal output achieved in this experiment occurred at the final readings

of the chemostat (0.48mW, compared to the previous maximum of 0.115mW) (Fig 8).

The corresponding current was 1.5mA. The resistance decreased even more, settling

at 220 ohms, emphasising that the bacteria just kept getting better. 0.002 W/m3 was

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the maximal volumetric power, also, being higher than the previous power (3.54x10 -5

W/m3).

Summing up, over time, it was clear to see that the bacteria improved their ability to

make electricity. The maximum voltage increased over the four set of results (from

0.272V to 0.556V) and the voltages stayed higher over lower resistances. The

resistance at which the power was maximal kept decreasing (2500 ohms to 220

ohms), also showing the improvement of bacteria over time (days/weeks). On top of

this, a continuous improvement in the maximal volumetric power (1.39E-05 W/m3 to

0.0019 W/m3) and maximal power (0.003474 mW to 0.477164 mW) was evident, too.

However, at a batch coulombic efficiency of only 0.665%, this shows that the

bacteria, in the batch culture, were extremely weak.

Results section could have been more structured by using sub-chapters.

Also the results section focuses just on

You did not mention how you judged that acetate was degraded.

Discussion

In this experiment, the behavioural characteristics of microbial fuel cell was

investigated and observed. From the results, it confirmed the hypothesis that bacteria

can be forced to act like a battery under certain conditions. The results evidently

showed that the bacteria were getting better or more capable of charging up the

electric potential in the microbial chamber. This can be observed from Figure 3 and 7,

showing a two fold increase in voltage readings from 0.272 V (first batch results) to

0.556 V (second chemostat results) at 1M ohms resistance.

Other than voltage, the MFC also recorded better readings in key performance

parameters, such as maximum power output at corresponding resistances. This is

apparent from Figure 4 and 8 where the maximum power output increased from

0.0035 mW at 2500 ohms (first batch results) to 0.48 mW at 220 ohms (second

chemostat results). The increase in maximum power output and voltage at decreasing

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resistance confirmed that the bacteria were improving in health; resulting in an

increase in the amount of bacteria which can transfer electrons to the carbon anode.

During the course of this experiment, the MFC ran in two feeding modes which were

fed-batch and chemostat batch. However, the chemostat mode of this experiment is

technically more of a semi-continuous batch mode. (True) The reason for the phrase

‘semi-continuous’ was because the inflow and outflow of the bacteria anode chamber,

though having the characteristics of a chemostat, was only semi-continuous as it had

an inflow of 10 mL substrate 4 hours 48 minutes interval. A true chemostat has a

continuous inflow and outflow. In the semi-continuous batch mode, the bacteria may

be experiencing starvation during the 4 hours 48 minutes interval. This semi-

continuous feeding schedule may affect the bacteria physiologically. The reason for

the long interval is due to the limitations introduced by the pumping system which

allows the slowest pumping rate of 10 mL per minute. The reason why 50 mL of 0.02

M of acetate was fed to the bacteria per day during the semi-continuous mode, was

due to the fact that during the experiment for the coulombic efficiency in batch mode,

the bacteria took more than 20 hours to fully metabolise 1 mL of 1 M acetate.

The coulombic efficiency (CE) for this MFC in batch mode was 0.665 %. This is

significantly lower than the usual CE published in literatures which range from 60-

80% [2]. From Figure 6, acetate oxidation rate was out by a factor of 111. This is

because, the lower the CE, the more inaccurate will the acetate oxidation rate be. The

low CE may be due to the fact that 1 mL of 1 M acetate was used in getting the feed

spike curve. This caused the long hours needed for the acetate added to be fully

metabolised, which may result in inaccuracies. A aliquot of 30 uL of 1 M acetate

should produce a more accurate result. The coulombic efficiency for the semi-

continuous mode was not done as it was impossible to determine the amount of

acetate present in the anode chamber at any one time without doing gas

chromatography. However, the results also clearly showed that the MFC in semi-

continuous batch mode was significantly more capable than the MFC in batch mode

as its power output and maximum potential build up were both almost twice of that in

batch mode. However, no further concrete conclusions can be made from this

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comparison as the biomass in both modes were not collected and measured. Further

investigation is recommended in determining the amount of biomass present in semi-

continuous batch and batch mode as it is expected that the amount of biomass in semi-

continuous batch will be significantly more than those in batch mode.

Further experiments should be conduced to find out whether the increase of potential

was caused by the increase in amount of bacteria or simply the bacteria were

becoming more efficient in donating electrons to the anode.

Hence, it is recommended for further experiments, the weight of the carbon sponge to

be weighed first before starting the experiment to allow the biomass to be measured.

The effect of temperature on the electric potential produced by the bacteria is worth

further investigations, as the increase in temperature to 30°C increases the electric

potential significantly. Changing different substances at the cathode chamber may

also produce different results worth investigating.

Overall, clearly written apart from some sections as indicated.

The only results you have shown and discussed is polarisation curves.

The feeding rate of the chemostat was not adjusted to the rate of current.

With this quality of writing and understanding the project would have been

excellent had it started earlier (allowing the buildup of biomass with

coulombic efficiencies higher than 20%.

Comparison of own reslts with literature or other authors missing.

Marks: 7/10 because of the reasonably good writing and good

understanding. More versatile experiments, more comparision with the

literature and better explanations of terms findings could have given more.

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References

Atkins, P. & De Paula, J. (2006) Atkins' Physical Chemistry, Oxford University Press.

Gangrekar M.M & Shinde V.B (2006) Performance of membrane-less microbial fuel

cell treating wastewater and effect of electrode distance and area on electricity

production. Bioresource Technology, 97, 543-550.

Ieropoulos, I., Greenman, J., Melhuish, C. & Hart, J. (2005) Comparative study of

three types of microbial fuel cell. Enzyme and Microbial technology, 37, 238-245.

Larminie, J. & Dicks, A. (2003) Fuel Cell Systems Explained, West Sussex, John

Wiley and Sons LTD.

Liu, H., Ramanarayanan, R. & Logan, B. (2004) Production of electricity during

wastewater treatment using a single chamber microbial fuel cell. Environmental

Science and Technology, 38, 2281-2285.

Min, B. & Logan, B. (2004) Continuous electricity generation from domestic

wastewater and organic substrates in a flat plate microbial fuel cell. Environmental

Science and Technology, 38, 5809-5814.

Mohan, S., Raghavulu, S., Srikanth, S. & Sarma, P. (2007) Bioelectricity production

by mediatorless microbial fuel cell under acidophillic condition using wastewater as a

substrate: Influence of substrate loading rate. Current Science, 92, 1720-1726.

Moon, H., Chang, I. S. & Kim, B. H. (2006) Continuous electricity production from

artificial wastewater using a mediator-less microbial fuel cell. Bioresource

Technology, 97, 621-627.

Serway, R. & Faughn, J. (2003) College Physics, Melbourne, Thompson Brooks\Cole.

Stams, A., De Bok, F., Plugge, C., Van eekert, M., Dolfing, J. & Schraa, G. (2006)

Exocellular electron transfer in anaerobic microbial communities. Environmental

Microbiology, 8, 371-382.

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Williams, K. (Ed.) (1966) An Introduction to Fuel Cells, London, Elsevier Publishing.

Recommendations (Overview)

Intrepid biotechnologists, congratulations on choosing the bioelectricity project. Here are a few of points to help you on your journey.

1 – The anode chamber is the brown sludgy one. The cathode chamber is the one that looks like cordial. Someone who marks your work will probably ask you that sometime, so try and remember it. If you can’t remember it, then leap forward keenly as if you are bursting with the answer, and Ralf will tell you to shutup and pick on someone else.

2 – The anode is not really anaerobic because the oxygen is bad for electricity production, it is because it stinks like real sewerage even though it is fake (at least they told me it was)

3 – The cathode solution is not very nice. I don’t think there is even any real cordial in there. Maybe the cyanide bit is a hint.

4 – Ralf likes pumps, so put one in there early on, even if it is just for looks. That way you will still get to do nothing for the first two weeks, but no-one will bug you about progress. It doesn’t hurt to keep him happy, and not many people are so easily amused.

5 – Don’t worry so much about polarisation curves. I think it will give someone a headache to mark anyway. Just put a pump on it and play around with lots of different things, (I agree!) I would suggest dilution rate and different oxidising agents in the cathode chamber. Measure the current which you achieve at steady state. It is quicker too.

If anyone actually figures out what a polarisation curve is, can you please email me?

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6 – Kayu is the most helpful, knowledgeable and genuinely kind demonstrator you will ever come across. Pay attention to this guru. Even though he is quiet he is packing the most powerful MFC in the world today in that lab of his. He will probably rule the world one day by solving the energy crisis with a car that runs on sewerage with the added bonus of not having to stop for toilet breaks on long trips.

Recommendations (In detail)

1. It is best to place the MFC into a tank of 30°C as early as possible. This is the

optimum temperature for the bacteria to live in.

2. Ensure the water level in the incubation tank is at the correct height level at all

times.

3. The setup should be done in such a way so that the wire connecting to the two

electrodes are not being disturbed too much. ie. Use crocodile clip wires in

between the electrodes and the resistor & voltmeter.

4. Current should be calculated using Ohm’s law instead of reading off the

voltmeter.

5. The MFC equipments are all very expensive. Handle with care.

6. It is recommended to go straight into chemostat mode and conduct

experiments in that mode instead of using precious time doing a batch mode.

7. A pump which can pump very small amounts of substrate for very short

intervals of time should be secured as early as possible for the experiment.

8. Stirring rate is recommended to be at around 200 rpm. Too high a stirring rate

may dislodge the electricity producing bacteria from the anode.

9. The total concentration of acetate in the anode chamber at any one time is

recommended to be not more than 0.1 mM to 0.5 mM.

10. After running the MFC for 3 to 5 days, the blackish sludge can be poured

away carefully and slowly and replace the anode chamber with synthetic

wastewater. Ensure that the carbon sponge anode is not disturbed too much

during the transition as the electric producing bacteria should be adhering

themselves to it. Initial voltage may be lowered drastically but will increase

dramatically soon after.

21

11. pH should be monitored frequently to ensure that the pH does not rise or fall

too much.

12. If carbon source was added into the synthetic wastewater, proper measures

should be taken to prevent or delay its contamination with bacteria in the

surroundings.

13. Ensure that the circuit is not opened for too long as the bacteria may die off

due to lack of means to transfer electrons.

14. If any stirring is to be introduced in any thing, ensure that it is as air tight as

possible to reduce oxygen dissolving into the system. The system is to be keep

as anaerobic as possible for maximum electricity production.

15. Ferricyanide should be changed in the cathode when it turns turbid or loses

colour. Alternatively, peroxide can be added to reoxidise it.

16. If a pump is added, a compromise in the air-tightness of the system may occur.

This is because an opening need to be introduce so that an inflow and outflow

can occur. Without the opening, the pressure in the vacuum will prevent the

pump from doing its job. Measures can be taken to both reduce the oxygen

dissolution as well as contamination.

17. Prior knowledge on MFC and electrochemistry is preferable to do this

experiment. The first week should be used to familiarise these concepts from

literature.

18. Using a polarisation and power curve is the best way to illustrate that the

bacteria culture in the anode chamber is getting enriched as the experiment

proceeds. Similarly, columbic efficiency should improve over time.

19. When producing the polarisation/power curves, there is no need to wait for the

voltage to completely stabilise. It could take half a day to a day to stabilise. As

soon as the increase in voltage, during the charging up, is significantly slow

(about 0.1 mV every 10 to 15 minutes) it is good enough to be considered as

stabilised. Proceed with the experiment from there.

22

20. Then doing the columbic efficiency curve, 30 uL of 1 M acetate solution is

more than enough to produce the “feed spike”. Adding more than that will

result in long hours of waiting time for all the added acetate to be fully

metabolised.

21. As this experiment is very new and you will probably be the second group to

be doing this, MAKE SURE you know exactly what your experiment is going

to achieve and how are you going to carry it out latest by 3rd week. This is

probably loads of things you can find out about MFCs but be realistic and

choose those that interest you most and discuss with your supervisors.

22. You will probably hear a lot of recommendations on what you can do with

your experiment. Listen to those BUT choose only those that you think you

are able to carry out. Time, equipment, complexity of the tasks should be some

of your considering factors.

23. During the first week, have a serious meeting and layout house rules. Identify

those members who are most likely not going to have much contribution and

lay down the terms and conditions. This is essential and should be done as

soon as possible.

Quite some useful hints here. Well done.

In summary, this project is relatively easy, and will probably be a lot easier for you

than for us. It will better if any group doing this project has a bit of Chemistry and

Physics background, don’t panic, just the basic stuff. You can get very technical on

this one, but I would suggest playing around with more things and collecting simpler

information, maybe columbic efficiency and current output at steady state. I think we

made a valuable contribution to science too. We probably generated enough power to

run a single LED to read by for 2 seconds. It is amazing what you can read in two

seconds. Other methods of power generation from sewerage might also be worth

investigating. Like putting it on your garden, growing potatoes, feeding them to rats,

and making the rats generate power on a treadmill. Try and put a pump in there

though to keep Ralf happy (BLESS HIM).

23

Good luck!

Acknowledgement

We would like to thank our

lecturer, Dr Ralf, for his valuable

guidance and advices for our

project.

We would like to thank our Mentor,

Kayu Cheng who assisted us in

configuring and the setting up of

Microbial Fuel Cell.

24

Appendix

Materials and equipment

1L Sodium acetate 1.0M for batch culture

1L Sodium acetate 0.02M for chemostat

1L activated sludge

2 electronic timers (LT48W)

2 magnetic stirrer bars

2000mL synthetic waste added with 2.mL of trace elements

3 Terumu needles (0.65 x 32mm) with syringes

70% ethanol spray

Aquarium (500mL)

Bandages (for insulation purpose)

Cotton wool (for insulation purpose)

Crocodile clips wire

Distilled water

Electronic speed adjustable pump (Chemap AG)

Esky box

Hot plate stirrer

25

MaCartney bottles

Oxygen probe (Hanna instrument HI9145)

Pen knife

pH meter

Pipette tips

Pipette (P200)

Potassium Ferricyanide ~ 50mM at pH7 + 100mL Phosphate biffer ~ 100mM

PVC fasteners

Trace element (1.25mL/L)

Voltmeter

Rubber pipes (approximately 1.5 meters for whole experiment)

Resistor

Scissor

1L Synthetic wastewater consists 20mL of sodium acetate 1.0M, 1.25mL trace

elements, 480mg NaHCO3, 95.5mg NH4Cl, 10.5mg K2HPO4, 5.25mg KH2PO4,

63.1mg CaCl2.2H20 and 19.2mg MgSO4.7H2O.

26

A set of MFC which consists of the following:

Figure 9: Batch culture set up

27

Figure 10: Chemostat set up

28

Figure 11: In detail of chemostat set up

29

Table 1: First set of results taken to form a power and polarisation curve.

Resistance (Ohms)

Voltage (V)

Voltage (mV)

Current (mA)

Power (mW)

Volumetric Power (W/m3)

1000000 0.272 272 0.000272 7.4E-05 2.96E-07560000 0.2724 272.4 0.000486 0.000133 5.3E-07330000 0.2725 272.5 0.000826 0.000225 9E-07220000 0.2724 272.4 0.001238 0.000337 1.35E-06100000 0.272 272 0.00272 0.00074 2.96E-0668000 0.2704 270.4 0.003976 0.001075 4.3E-0647000 0.2688 268.8 0.005719 0.001537 6.15E-0633000 0.2664 266.4 0.008073 0.002151 8.6E-064700 0.1205 120.5 0.025638 0.003089 1.24E-052500 0.0932 93.2 0.03728 0.003474 1.39E-051150 0.0618 61.8 0.053739 0.003321 1.33E-051000 0.055 55 0.055 0.003025 1.21E-05660 0.0386 38.6 0.058485 0.002258 9.03E-06550 0.033 33 0.06 0.00198 7.92E-06330 0.0218 21.8 0.066061 0.00144 5.76E-06150 0.0103 10.3 0.068667 0.000707 2.83E-0656 0.0054 5.4 0.096429 0.000521 2.08E-0633 0.0032 3.2 0.09697 0.00031 1.24E-065 0.00049 0.49 0.098 4.8E-05 1.92E-07

Table 2: Second set of results taken to form a power and polarisation curve

Resistance (Ohms)

Voltage (V)

Voltage (mV)

Current (mA)

Power (mW)

Volumetric Power (W/m3)

1000000 0.278 278 0.000278 7.73E-05 3.09E-07560000 0.277 277 0.000495 0.000137 5.48E-07330000 0.277 277 0.000839 0.000233 9.3E-07220000 0.277 277 0.001259 0.000349 1.4E-06100000 0.276 276 0.00276 0.000762 3.05E-0668000 0.274 274 0.004029 0.001104 4.42E-0647000 0.272 272 0.005787 0.001574 6.3E-0633000 0.2703 270.3 0.008191 0.002214 8.86E-0622000 0.2615 261.5 0.011886 0.003108 1.24E-0515000 0.2544 254.4 0.01696 0.004315 1.73E-0510000 0.2438 243.8 0.02438 0.005944 2.38E-056800 0.2268 226.8 0.033353 0.007564 3.03E-054700 0.2025 202.5 0.043085 0.008725 3.49E-052200 0.1396 139.6 0.063455 0.008858 3.54E-051500 0.1074 107.4 0.0716 0.00769 3.08E-051000 0.0796 79.6 0.0796 0.006336 2.53E-05

30

820 0.0682 68.2 0.083171 0.005672 2.27E-05330 0.0308 30.8 0.093333 0.002875 1.15E-05150 0.0145 14.5 0.096667 0.001402 5.61E-0656 0.0055 5.5 0.098214 0.00055 2.16E-0633 0.0034 3.4 0.10303 0.00035 1.4E-0610 0.0011 1.1 0.11 0.000121 4.84E-075 0.00006 0.6 0.12 0.000072 2.88E-07

Table 3: First set of chemostat results taken to form a power and polarisation curve

Resistance (Ohms)

Voltage (V)

Voltage (mV)

Current (mA)

Power (mW)

Volumetric Power (W/m3)

1000000 0.453 453 0.000453 0.000205 8.21E-07560000 0.453 453 0.000809 0.000366 1.47E-06330000 0.453 453 0.001373 0.000622 2.49E-06150000 0.453 453 0.00302 0.001368 5.47E-0682000 0.452 452 0.005512 0.002492 9.97E-0647000 0.453 453 0.009638 0.004366 1.75E-0510000 0.437 437 0.0437 0.019097 7.64E-054700 0.412 412 0.08766 0.036116 0.0001441000 0.308 308 0.308 0.094864 0.000379560 0.254 254 0.453571 0.115207 0.000461220 0.136 136 0.618182 0.084073 0.000336100 0.0634 63.4 0.634 0.040196 0.00016147 0.0335 33.5 0.712766 0.023878 9.55E-0522 0.016 16 0.727273 0.011636 4.65E-055 0.0037 3.7 0.74 0.002738 1.1E-05

Table 4: Second set of chemostat results taken to form a power and polarisation curve

Resistance (Ohms)

Voltage (V)

Voltage (mV)

Current (mA)

Power (mW)

Volumetric Power (W/m3)

1000000 0.556 556 0.000556 0.000309 1.23654E-06560000 0.556 556 0.000993 0.000552 2.20811E-06330000 0.556 556 0.001685 0.000937 3.7471E-06150000 0.556 556 0.003707 0.002061 8.24363E-0682000 0.557 557 0.006793 0.003784 1.51341E-0547000 0.557 557 0.011851 0.006601 2.64042E-0510000 0.548 548 0.0548 0.03003 0.0001201224700 0.535 535 0.11383 0.060899 0.0002435961000 0.473 473 0.473 0.223729 0.000894916560 0.428 428 0.764286 0.327114 0.001308457

31

220 0.324 324 1.472727 0.477164 0.001908655100 0.215 215 2.15 0.46225 0.00184947 0.105 105 2.234043 0.234574 0.00093829822 0.052 52 2.363636 0.122909 0.0004916365 0.012 12 2.4 0.0288 0.0001152

32

Table 5: Calculations for Columbic Efficiency.

Time Normalized Curr Mean Curr

interv. voltage Resistance Current with baseline curr in intrv. Coulomb

sec sec mV Ohm mA mC/s mC/s mC/s mC C

0 61.5 999 0.06156 0.06156 0.0000

3600 3600 61.5 1000 0.0615 0.0615 -0.0001 0.062 222 0.222

4500 900 61.8 1000 0.0618 0.0618 0.0002 0.062 55 0.055

4920 420 62.1 1000 0.0621 0.0621 0.0005 0.062 26 0.026

5520 600 62.3 1000 0.0623 0.0623 0.0007 0.062 37 0.037

6120 600 62.6 1000 0.0626 0.0626 0.0010 0.062 37 0.037

6720 600 62.8 1000 0.0628 0.0628 0.0012 0.063 38 0.038

7320 600 63 1000 0.063 0.063 0.0014 0.063 38 0.038

7920 600 63.2 1000 0.0632 0.0632 0.0016 0.063 38 0.038

8520 600 63.4 1000 0.0634 0.0634 0.0018 0.063 38 0.038

9120 600 63.5 1000 0.0635 0.0635 0.0019 0.063 38 0.038

9720 600 63.6 1000 0.0636 0.0636 0.0020 0.064 38 0.038

12720 3000 65.3 1000 0.0653 0.0653 0.0037 0.064 193 0.193

33

13320 600 65.7 1000 0.0657 0.0657 0.0041 0.066 39 0.039

16320 3000 68.1 1000 0.0681 0.0681 0.0065 0.067 201 0.201

16920 600 68.4 1000 0.0684 0.0684 0.0068 0.068 41 0.041

17520 600 68.9 1000 0.0689 0.0689 0.0073 0.069 41 0.041

18120 600 69.4 1000 0.0694 0.0694 0.0078 0.069 41 0.041

18720 600 69.7 1000 0.0697 0.0697 0.0081 0.070 42 0.042

20520 1800 71 1000 0.071 0.071 0.0094 0.070 127 0.127

24120 3600 72.6 1000 0.0726 0.0726 0.0110 0.072 258 0.258

24720 600 72.65 1000 0.07265 0.07265 0.0111 0.073 44 0.044

25920 1200 73 1000 0.073 0.073 0.0114 0.073 87 0.087

26520 600 73 1000 0.073 0.073 0.0114 0.073 44 0.044

27120 600 72.9 1000 0.0729 0.0729 0.0113 0.073 44 0.044

27720 600 72.8 1000 0.0728 0.0728 0.0112 0.073 44 0.044

28320 600 72.7 1000 0.0727 0.0727 0.0111 0.073 44 0.044

28920 600 72.6 1000 0.0726 0.0726 0.0110 0.073 44 0.044

29520 600 72.5 1000 0.0725 0.0725 0.0109 0.073 44 0.044

30120 600 72.4 1000 0.0724 0.0724 0.0108 0.072 43 0.043

34

30720 600 72.3 1000 0.0723 0.0723 0.0107 0.072 43 0.043

31320 600 72.2 1000 0.0722 0.0722 0.0106 0.072 43 0.043

31920 600 72.1 1000 0.0721 0.0721 0.0105 0.072 43 0.043

32520 600 72 1000 0.072 0.072 0.0104 0.072 43 0.043

33120 600 71.8 1000 0.0718 0.0718 0.0102 0.072 43 0.043

33720 600 71.7 1000 0.0717 0.0717 0.0101 0.072 43 0.043

34320 600 71.5 1000 0.0715 0.0715 0.0099 0.072 43 0.043

34920 600 71.4 1000 0.0714 0.0714 0.0098 0.071 43 0.043

35520 600 71.2 1000 0.0712 0.0712 0.0096 0.071 43 0.043

36120 600 70.3 1000 0.0703 0.0703 0.0087 0.071 42 0.042

36720 600 70 1000 0.07 0.07 0.0084 0.070 42 0.042

37320 600 69.6 1000 0.0696 0.0696 0.0080 0.070 42 0.042

60120 22800 65 1000 0.065 0.065 0.0034 0.067 1534 1.534

72120 12000 62.1 1000 0.0621 0.0621 0.0005 0.064 763 0.763

72720 600 62 1000 0.062 0.062 0.0004 0.062 37 0.037

73320 600 61.8 1000 0.0618 0.0618 0.0002 0.062 37 0.037

73920 600 61.7 1000 0.0617 0.0617 0.0001 0.062 37 0.037

35

74520 600 61.5 1000 0.0615 0.0615 -0.0001 0.062 37 0.037

75120 600 61.4 1000 0.0614 0.0614 -0.0002 0.061 37 0.037

75720 600 61.3 1000 0.0613 0.0613 -0.0003 0.061 37 0.037

76320 600 61.3 1000 0.0613 0.0613 -0.0003 0.061 37 0.037

76920 600 61.3 1000 0.0613 0.0613 -0.0003 0.061 37 0.037

Sum of C 5.1

36

37