1.

INTRODUCTION

1.1. Preface

Microbial fuel cells (MFC) have gained importance in the last few decades due to
their ability to produce energy, either as electricity or hydrogen, from renewable resources
such as waste water. Progress in design has given numerous aspects that are to be
investigated so as to attain a cell that may produce significant yields. In nature, all living
beings utilize some material as a source of food which is mandatory for deriving energy for
their sustenance and proliferation. Microorganisms such as bacteria utilize organic substances
inclusive of carbohydrates and alcohols to attain the required amount of energy. This intricate
process involves the degradation of the carbohydrate along the Tri-Carboxylic Acid cycle. In
the course of this process, produced are a good amount of electrons and protons. These
electrons find their way through the electron transport chain to combine with the natural
terminal electron acceptor, oxygen and forms water with the proton molecules.

A concept was devised and put to test where in the effect of replacing the natural
terminal acceptor, oxygen, with a substance that could utilize these electrons to generate
energy was stipulated. Thus began a succession of scientific study to develop such
technology that might exploit these naturally produced electrons for energy generation.

1.2. History

The idea of using microbial fuel cells in an attempt to produce electricity was first
conceived by M. C. Potter in 1911. A professor of botany at the University of Durham,
managed to generate electricity from E. coli. In 1931, Barnet Cohen drew more attention to
the area when he created a number of microbial half fuel cells that, when connected in series,
were capable of producing over 35 volts, though only with a current of 2 milliamps. More
work on the subject came with a study using hydrogen produced by the fermentation of
glucose by Clostridium butyricum as the reactant at the anode of a hydrogen and air fuel cell.
Unfortunately, though the cell functioned, it was found to be unreliable due to the unstable
nature of the hydrogen production from the micro-organisms. This issue was later resolved by
Karube in 1976, when he proposed the current design concept of an MFC which came into
existence a year later.
 Even by the time of Suzuki’s work in the late seventies, little was understood about
how these microbial fuel cells functioned. The idea was picked up and studied later in more
detail first by MJ Allen and then by H. Peter Bennetto both from King’s College London.
Bennetto saw the fuel cell as a possible method for the generation of electricity for third
world countries. His work, starting in the early 1980s helped to build an understanding of
how fuel cells operate and until his retirement was seen by many as the foremost authority on
the subject.

In May 2007, the University of Queensland, Australia, completed its prototype MFC,
as a cooperative effort with Fosters Brewing Company. The prototype, a 10 liter design,
converts the brewery waste water into carbon dioxide, clean water, and electricity. With the
prototype proven successful, plans are in effect to produce a 660 gallon version for the
brewery, which is estimated to produce 2 kilowatts of power.

1.3. Indian Scenario

The energy crisis in India has reached at alarming levels requiring immediate
attention. With burgeoning population, it is vital to find alternative methods of energy
generation, organic waste utilization that are sustainable for future. Energy consumption
increased dramatically and an unbalanced energy management exists. While there is no sign
that this growth in demand will abate, particularly amongst the developing nations like India,
there is now an awareness of the transience of nonrenewable resources and the irreversible
damage caused to the environment.

Bio-fuel cells potentially offer solutions to all these problems, by taking nature’s
solutions to energy generation and tailoring them to our own needs.

1.4. World scenario

The need for an alternate fuel is such that extensive research is under progress in
identifying a potential method and substrate for its application in energy production. The
building of a sustainable society will require reduction of dependency on fossil fuels and
lowering of the amount of pollution that is generated. Waste treatment is an area in which
these two goals can be addressed simultaneously. As a result, there has been a paradigm shift
recently, from disposing of waste to using it. Waste from household, industries and
agriculture are ideal candidates of substrates for energy generation because they contain high
levels of easily degradable organic material.

The building of sustainable society requires reduction in dependence of fossil fuel and
lowering the amount of pollution that is generated. Wastewater treatment is an area in which
these two goals can be addressed simultaneously. Wastes from households, industries etc are
ideal candidates of substrates for energy generation because they contain a high amount of
easily degradable organic matter.

1.5. Biomass- A potential source

Biomass is ubiquitous in nature and has immense potential as a substrate for energy
production. Ethanol, Butanol, Methanol, Methane, Biodiesel, Biohydrogen and Bioelectricity
could be produced from biomass. In India, production of Gobar gas by anaerobic digestion
has been successfully carried out, but its application is limited to only rural areas. It also has
issues related to sustainability and environmental problems.

The production of energy from renewable substrates, such as biomass, is important for
creating sustainable energy production and reducing global emissions of CO2. Hydrogen can
be an important component of an energy infrastructure that reduces CO 2 emissions, if
hydrogen is produced from non-fossil fuel sources and used in fuel cells. Hydrogen gas can
be biologically produced at high concentration (60%) from the fermentation of high sugar
substrates such as glucose and sucrose. However, known fermentation routes can produce
only 33% of the maximum potential energy from a sugar such as glucose. More commonly,
yields of only half this amount are achieved resulting in the remainder of the energy
(typically 85%) being tied up in non-fermentable or poorly fermented organic acids and
solvents such as acetic acid, butyric and propionic acids, ethanol, and Butanol (Logan et al.,
2002).

Instead of producing electricity indirectly from organic materials with biologically-

generated hydrogen, it is now known that electricity can be produced directly from the
degradation of organic matter in a MFC (Suzuki et al., 1978; Allen and Bennetto, 1993; Kim
et al., 2002; Bond and Lovley, 2003; Liu et al., 2004; Liu and Logan, 2004; Oh et al., 2004).
1.6.1. Fuel Cells

A fuel cell is an electrochemical device that continuously converts chemical energy to

electrical energy as long as fuel and oxidants are supplied to it. A Biological fuel cell may be
defined as an assembly of an electrochemical device that converts chemical energy derived
from a biological substrate such as carbohydrates like sugar and alcohol directly into
electrical energy.

With the advent of space research, concept of fuel cell was introduced. The need for a
miniature device necessitated the fabrication of fuel cell to meet the requirement of power in
a short span of time. The chemical fuel cell fabrication involves high cost chemicals and
extreme operational conditions. Enzymatic fuel cells are much beyond the purview of
commercialization due to the cost involved in enzyme production and purification.

There are two types of biological fuel cells, namely microbial fuel cells and enzymatic
fuel cells.

1.6.2. Microbiological Fuel Cell

The use of microorganisms in biological fuel cells eliminates the isolation of

individual enzymes, thereby providing cheaper substrates for biological fuel cells.
Microorganisms can be used in four ways for generating electrical energy:

Microorganisms can produce electrochemically active substances through

fermentation or metabolism. For the purpose of energy generation, fuels are produced in
separate reactors and transported to the anode of a conventional fuel cell. Accordingly, in this
configuration, the microbial reactor is kept separated from the fuel cell.

In the second configuration, the microbiological fermentation process proceeds

directly in the anodic compartment of the fuel cell.

In the third configuration, electron-transfer mediators shuttle electrons between the

microbial biocatalytic system and the electrode. The mediator molecules accept electrons
from the biological electron transport chain of the microorganisms and transport them to the
anode of the biological fuel cell.
 In the fourth configuration, the metal reducing bacterium having cytochromes in its
outer membrane and their ability to communicate electrically with the electrode surface
directly results in a mediator-less biological fuel cell.

1.6.3. Enzymatic fuel cells

Enzymes have a complex structure comprising proteins. The electron-transferring unit

of the enzyme, namely the apoenzyme and cofactor, is deeply buried inside its complex
structure. Hence, efficient electrical communication between the electrode substrate and the
enzyme biocatalysts is difficult. Mediated bio electrolysis uses small redox molecules as
electron transfer mediators between the enzyme and the electrode. This is useful in studying
electron transfer kinetics of the enzymes.

1.6.4. Microbial Fuel Cells

Microbial Fuel Cell is a device designed for a novel purpose of electricity generation
in the process of waste water treatment. Hence, it is an ideal solution for sustainable non
renewable source of energy. MFC can be best defined as a fuel cell where microbes act as
catalyst in degrading the organic content to generate electricity. MFCs operate by directly
capturing the electrons generated when electrochemically active bacteria breakdown organic
substrates. The existence of a potential gradient across the microbial cell membrane forms the
basis for the generation of bioelectricity.

Various design modifications employed throughout these years have given significant
yields and opened up a new frontier of research. MFC involves a multidisciplinary approach
and this study focuses on the biological, design, physical and chemical parameters involved
in MFC. Microbially catalyzed electron liberation at the anode and subsequent electron
consumption at the cathode, are the defining characteristics of an MFC.

It has been shown that certain metal reducing bacteria, belonging to the family
Geobacteraceae, can directly transfer electrons to the electrodes using electrochemically
active redox enzymes such as cytochromes present in the outer membrane. These cells that do
not employ additional chemical substances called mediators, have come to be known as
mediator less MFCs and have more commercial applications compared to their counterparts,
as the mediator used in the biofuel cells, are expensive and potentially toxic to
microorganisms growing in the substrate.
1.6.5. Proton Exchange Membrane fuel cells

Although fuel cells have been used since the 1960s as power supplies for satellites,
recently renewed attention for their potential to efficiently produce clean energy from
hydrogen has increased. Nafion was found effective as a membrane for proton exchange
membrane fuel cells by permitting hydrogen ion transport while preventing electron
conduction. Solid Polymer Electrolytes, which are made by connecting or depositing
electrodes (usually noble metal) to both sides of the membrane, conduct the electrons through
an energy requiring process and rejoin the hydrogen ions to react with oxygen and produce
water. Fuel cells are expected to find strong use in the transportation industry.

Nafion is a sulfonated tetrafluoroethylene copolymer discovered in the late 1960s by

Walther Grot of DuPont de Nemours. It is the first of a class of synthetic polymers with ionic
properties which are called ionomers. Nafion's unique ionic properties are the result of
incorporating perfluorovinyl ether groups terminated with sulfonate groups onto a
tetrafluoroethylene (Teflon) backbone. Nafion has received a considerable amount of
attention as a proton conductor for proton exchange membrane fuel cells because of its
excellent thermal and mechanical stability. Microbial fuel cells are typically designed as a
two-chamber system with the bacteria in the anode chamber separated from the cathode
chamber by a polymeric proton exchange membrane (Gottesfeld et al., 1997).

1.7. Substrate for MFC

The substance that serves efficiently as a substrate in MFC must essentially harbour
optimum populations of microbial colonies that effectively bring about greater movement of
electrons in the circuit and hence must it be abundant in those constituents that these
microorganisms would find essential for their survival and metabolic livelihood. If specific
populations of microbial strain in a special consortium were to bring in to effect a greater
deflection in the multimeter, the substrate should be able to provide a suitable environment,
nutrients and all other specifics that would critically maintain this consortium, and put into
effect this proposed increase in the electricity production. The substrate must also be flexible
and adaptable to changes made in any ancillary components added to the substrate in order to
modify the medium, ecosystem to favor the growth of specific populations that are found to
produce better result in the MFC. Waste water harbors varied range of microorganisms,
which include aerobes and anaerobes. Also with time its composition varies and thus the type
and number of organisms will fluctuate. A wide variety of fungi, protozoa, algae, bacteria and
virus are present in the waste water. Given these particular justifications, we have attempted
to produce electricity in this particular design of the MFC using waste water collected from a
domestic waste water treatment plant as a substrate of growth of the required
microorganisms.

1.8. Working of Fuel Cell

A MFC ordinarily consists of two chambers, one anaerobic (anode) and the other
aerobic (cathode). In the anaerobic chamber, substrate is oxidized by bacteria and the
electrons transferred to the anode either by an exogenous electron carrier or mediator or
directly from the bacterial respiratory enzyme to the electrode. In the latter case, the MFC is
known as a mediator-less MFC (Kim et al., 1999, 2002; Bond and Lovley, 2003; Gil et al.,
2003; Chaudhuri and Lovley, 2003; Rabaey et al., 2003; Jang et al., 2004). The anaerobic
chamber is connected internally to the aerobic chamber by a proton-conducting material, and
externally by a wire that completes the circuit. In the aerobic chamber, electrons that pass
along the circuit combine with protons and oxygen to form water. Mediator-less MFCs have
the potential to produce electricity from anaerobic sediments for marine devices (Reimers et
al., 2001; Bond and Lovley, 2003) and electricity from sewage (Gil et al.,2003; Liu et al.,
2004). Mediator-less MFCs have only recently been developed, and therefore the factors that
affect optimum operation, such as the bacteria used in the system, the type of proton
conductive material, and the system configuration, are not well understood.

As most organic substrates undergo combustion with an evolution of energy, bio-

catalyzed oxidation of organic substances by oxygen at the two electrode interfaces
provides means for conversion of chemical energy to electrical energy. Through natural
cycles of metabolism, electrons produced in the microorganisms are donated to the anode
of the fuel cell under the anaerobic conditions, (absence of oxygen – terminal electron
accepter in the respiratory chain) the movement of these electrons through the circuit
containing the cathode thus results in a deflection in the multimeter used to detect and
measure the electricity produced.

Also produced in the substrate medium, by natural metabolism are H+ atoms that
naturally would have formed water in the presence of oxygen but the anaerobic conditions
would have to be moved out from the medium to avoid reduction in electricity production
by deflection of electrons. Hence these protons are moved to the cathode of the cell,
through a salt bridge or a proton exchange membrane (PEM) to form water. An aerobic
chamber enclosing the receptive cathode provides a suitable environment for this reaction.

In normal microbial catabolism a substrate such as carbohydrate is oxidized initially

without the participation of oxygen, while its electrons are taken up by an enzyme active –
site which acts as a reduced intermediate described as follows:

C6H12O6 + 6H2O Æ 6CO2 + 24H+ + 24 e- [E0 = 0.014v] (1)

In the absence of oxygen, the electrons are diverted to the electrode, pass through the outer
circuit and ultimately combine with an electron sink, namely, molecular oxygen as follows:

6O2 + 24H+ + 24 e- Æ 12H2O [E0 = 1.23v] (2)

When micro-organisms consume a substrate such as sugar (lactose) in aerobic

conditions they produce carbon dioxide and water, however when oxygen is not present they
produce carbon dioxide, protons and electrons as described below:

C12H22O11 + 13H2O ---> 12CO2 + 48H+ + 48e- (3)

1.9. Mediators

Microbial fuel cells use inorganic mediators to tap into the electron transport chain of
cells and steal these electrons that are produced. The mediator crosses the outer cell lipid
membranes and plasma wall; it then begins to liberate electrons from the electron transport
chain that would normally be taken up by oxygen or other intermediates. The now reduced
mediator exits the cell laden with electrons that it shuttles to an electrode where it deposits
them; this electrode becomes the electro-generic anode (negatively charged electrode). The
release of the electrons means that the mediator returns to its original oxidized state ready to
repeat the process. It is important to note that this can only happen under anaerobic
conditions, if oxygen is present then it will collect all the electrons as it has greater electro
negativity than the mediator.

1.10. Mediator less Microbial Fuel cells

Mediator-less microbial fuel cells have been engineered at the Korea Institute of
Science and Technology, by a team led by Kim, Byung Hong year. A mediator-less microbial
fuel cell does not require a mediator but uses electrochemically active bacteria to transfer
electrons to the electrode (electrons are carried directly from the bacterial respiratory enzyme
to the electrode). Among the electrochemically active bacteria are, Shewanella putrefaciens
(Kim et al., 1999), Aeromonas hydrophila and others. Some bacteria, which have pili on their
external membrane, are able to transfer their electron production via these pili.