2.2 Basic Biological Processes in 2.

2 Basic Biological Processes in

Prokaryotes and Eukaryotes Prokaryotes and Eukaryotes
Objectives
At the end of this lesson, student should be able
to:
Describes the process of photosynthesis (Calvin
cycle) cycle)
Differentiate aerobic respiration (Krebs cycle)
with anaerobic respiration (various terminal
electron acceptors)
Evaluate the factors and condition to enhance
microbial growth kinetics
2.2 Basic Biological Processes in 2.2 Basic Biological Processes in
Prokaryotes and Eukaryotes Prokaryotes and Eukaryotes
A biological process is a process of a living organism.
Biological processes are made up of any number of
chemical reactions or other events that results in a
transformation.
May include:
Cell adhesion and aggregation Cell adhesion and aggregation
Cell communication / signaling
Morphogenesis (shapes of tissues, organs and entire
organisms)
Cell physiology
Reproduction / microbial kinetics
Interaction between organisms (population dynamics)
2.2.1 Cell Physiology 2.2.1 Cell Physiology
Physiology (from Greek: physis = nature, origin;
and logos = word, speech; lit. "to talk about the
nature (of things)") is the study of the mechanical,
physical, and biochemical functions of living
organisms. organisms.
Here we shall pay particular focus on biochemistry
and abiotic growth conditions. If we have some
understanding of the chemical basis of life, it will
help in our understanding of the way in which
microorganisms function, how they can be involved
in biogeochemical cycles but also how we can
make use of them in treating water and wastewater
Taxonomy of cells Taxonomy of cells
Taxonomy is the system of nomenclature which
biologists use to identify living things. Under this
system all living things are classified into the
following sets and subsets:
Kingdom Kingdom
Phylum
Class
Order
Family
Genus
Species
Taxonomy of cells Taxonomy of cells
Based on 16S rRNA studies organisms have been
divided into six kingdoms:
Plants,
Animals,
Protista Protista
Fungi
Archaea
Bacteria.
2.2.2 Environmental Growth 2.2.2 Environmental Growth
Conditions Conditions Conditions Conditions
Carbon & Energy Requirements Carbon & Energy Requirements
Taxonomy is not a particularly useful way of viewing
these organisms if we are to make use of them in
engineering processes!
The classification according to carbon and energy
requirements outlined below is much more useful
Mode of Nutrition Energy Source Carbon Source
Photoautotroph Light CO
2
Chemoautotroph Inorganic chemicals CO
2
Photoheterotroph Light Organic compounds
Chemoheterotroph Organic compounds Organic compounds
Exercise: Select the groups of micro-organisms that can use the following
compounds as a carbon or energy source.
a) Glucose; b) SO
4
; c) chlorophyll
Nutrient Requirements Nutrient Requirements
All micro-organisms need O, C, N, H, P and S, which
are the elements of organic matter.
They also need Na, K, Mg, Ca and Cl.
Trace elements that might be required include Mn,
Fe, Co, Cu, Zn, B, Al, V, Mo, I, Si, Sn, Ni, Cr, F and Fe, Co, Cu, Zn, B, Al, V, Mo, I, Si, Sn, Ni, Cr, F and
Se
Some micro-organisms require special growth factors
such as vitamins, amino acids etc.
Abiotic factors such as pH, presence and
concentrations of anions, temperature and pressure
can have important effects on bioavailability and
concentration of nutrients
Temperature & pH Range Temperature & pH Range
Different microorganisms have different temperature
and pH profiles and optimum T and pH for growth.
Oxygen Requirements Oxygen Requirements
Micro-organisms differ in their requirements for
oxygen. Some are:
Obligate aerobes
Obligate anaerobes
Facultative aerobes / anaerobes Facultative aerobes / anaerobes
Water Requirements Water Requirements
All microorganism need to be in an environment
where water is present in order to grow. Some fungi
can grow in relatively dry conditions on solid media
but the atmosphere must be damp.
Solutes (sugar, salts, etc.) dissolved in water have an
affinity for water, and this is therefore unavailable to affinity for water, and this is therefore unavailable to
micro-organisms. Therefore the more dissolved salts
the less likely it is that microorganisms will survive.
The availability of water can be expressed as water
activity, which is calculated as the ratio of the vapour
pressure of a solution divided by the vapour pressure
of pure water at the same temperature.
2.2.3 From Microbiology to 2.2.3 From Microbiology to
Biochemistry Biochemistry
or or or or
The Study of Metabolical Pathways in Microbial The Study of Metabolical Pathways in Microbial
Cells Cells
Cells and Chemicals Cells and Chemicals
In the previous section we considered the function of
living organisms at the level of the cell which is a
closed system in thermodynamic / mass balance
terms.
In cell biology we look at the sub-cellular components In cell biology we look at the sub-cellular components
and in biochemistry we consider the chemicals that
make up life.
Hierarchies based on size and complexity of the
molecules are shown in the next slide
Cells and Chemicals Cells and Chemicals
Cell
Organelles
Macromolecules
Chemical
Mitochondria
Nucleus
Ribosomes
Proteins, Lipids,
Polysacch., Nucleic Acid
Relationship between the cell and chemicals
Monomers
Organic Chemicals
Precursor Molecules
Polysacch., Nucleic Acid
Amino Acids, Nucleotides,
Sugars, Fatty Acids
e.g. Acetate, alcohols
e.g. H
2
, O, CO
2
, N
2
Metabolism Metabolism
Chemical Reactions
Within a living cell, there is a constant turn-over of
biochemicals (carbohydrates, lipids, proteins, nucleic acids
and combinations thereof) in many hundreds, possibly
thousands, of enzyme-catalysed chemical reactions. This is
metabolism, i.e. all the chemical reactions that occur within
a cell or organism. a cell or organism.
Metabolism has many different sets of chemical reactions
known as metabolic pathways. In order to simplify this
complex situation, two major types of pathways can be
considered:
Catabolism is the breakdown of large molecules into small ones to
release energy.
Anabolism is the building-up of small molecules to produce large
molecules, which requires energy.
Metabolism Metabolism
Catabolism and anabolism are related as shown in the figure. During
anabolism, adenosine triphosphate (ATP) is broken down to ADP and
inorganic phosphate, liberating energy that can then be used in the
synthesis of biochemicals.
Metabolism Metabolism
A more detailed study of the
reactions involved in catabolism
reveals that there are a number of
points at which ATP is produced.
These are identified in the figure for
the aerobic respiration of proteins,
carbohydrates and lipids. In aerobic
respiration, the micro-organisms use respiration, the micro-organisms use
oxygen as the ultimate electron
acceptor in catabolism.
Anaerobic respiration is also
possible when certain microbial
groups are capable of using
oxidised inorganic ions (nitrate,
sulphate, carbonate) instead of
oxygen as terminal electron
acceptor.
Photosynthesis
Converting light energy to chemical energy and storing it
in the bonds of sugar
Krebs or citric acid cycle
Anaerobic respiration
Using electron acceptors other than oxygen
Respiration without oxygen
Important in global nitrogen, sulphur and carbon
cycle cycle
Reduction process by microorganism
Aerobic respiration by nitrifiers Aerobic respiration by nitrifiers
Nitrifying organisms are chemoautotrophs and use carbon dioxide as their
carbon source for growth.
Nitrosomonas and Nitrosococcus oxidise ammonia with oxygen into nitrite
while the second step (oxidation of nitrite into nitrate) is (mainly) done by
bacteria of the genus Nitrobacter.
1
st
Step: NH
3
+ O
2
NO
2

+ 3H
+
+ 2e

2
nd
Step: NO
2

+ H
2
O NO
3

+ 2H
+
+ 2e

2 2 3
Exercise:
Discuss the rationale behind adding a nitrification inhibitor to BOD
5
samples as recommended by standard methods such as APHA , EPA.
Anoxic respiration by denitrifiers Anoxic respiration by denitrifiers
The process is performed primarily by heterotrophic bacteria (such as
Paracoccus denitrificans and various Pseudomonads) although
autotrophic denitrifiers have also been identified (e.g. Thiobacillus
denitrificans).
Denitrification only takes place in environments where oxygen
consumption exceeds the rate of oxygen supply such as in some soils
and groundwater, wetlands (e.g. Tasik Bera), poorly ventilated corners of
the ocean, and in seafloor sediments. Under these conditions bacteria the ocean, and in seafloor sediments. Under these conditions bacteria
respire nitrate as a terminal electron acceptor.
Denitrification proceeds through some combination of the following steps:
NO
3
-
NO
2
-
NO N
2
O N
2
(g)
Denitrification is the second step in the nitrification-denitrification process,
the conventional way to remove nitrogen from sewage and municipal
wastewater.
Fermentation Fermentation
Fermentation occurs during which energy is derived from the oxidation
of organic compounds, such as carbohydrates, using an endogenous
electron acceptor, which is usually an organic compound.
Note that fermentation can take place under aerobic or anaerobic
conditions!
The actual biochemical pathway the
reaction takes varies depending on reaction takes varies depending on
the sugars involved, but the most
common involves part of the glycolysis
pathway, which is shared with the
early stages of aerobic respiration in
most organisms. The later stages of
the pathway vary considerably
depending on the final product.
2.2.4 Microbial Growth Kinetics 2.2.4 Microbial Growth Kinetics
Introduction Introduction
Effective control of any system using biological processes is based on
an understanding of the basic principles governing the growth of micro-
organisms.
We have already looked at the environmental conditions, which have an
important effect on the survival of micro-organisms, like pH,
temperature, oxygen, water and nutrient and energy requirements. temperature, oxygen, water and nutrient and energy requirements.
Now we shall begin to look at the growth of micro-organisms and in
particular the kinetics of bacterial growth which is a key parameter in
the design of biological treatment processes.
Growth in batch culture Growth in batch culture
Bacteria reproduce by binary fission, that
is one cell simply divides into two.
The growth of a bacterial culture can be
represented by a curve that consists of
four stages or phases:
Lag phase - growth and reproduction are
just beginning
Log phase - reproduction is occurring at an
exponential rate
Stationary phase - environmental
surroundings and food supply cannot
support any more exponential growth
Death phase - when all of the nutrients
have been exhausted, the population dies
off
Specific growth rate Specific growth rate
In a batch reactor where there is no nutrient limitation or depletion and no
loss of cells due to endogenous metabolism or death, growth of biomass
will be exponential during the unlimited growth phase.
The specific growth rate is defined as the quantity of new biomass formed
per unit of original biomass per unit of time. Following equation can be
used:
t
t
t
e X X
t t
X
X

0
0 1
0
ln
X
t
Population at time t, e.g. number of cells per mL
X
0
Population at time zero
t time [hr]
specific growth rate [hr
-1
]
0 1 2 3 4 5 6 Time [hr]
Doubling time Doubling time
0 1 2 3 4 5 6 Time [hr]
The doubling time, t
d
, is analogous to the half-life for first order decay. It is The doubling time, t
d
, is analogous to the half-life for first order decay. It is
defined as the time taken for the population to double in size and is related
to the specific growth rate. X
t
= 2X
0

2 ln
2
ln ln
0
0
0
=

=
X
X
X
X
t
t
d
Exercise:
Calculate the maximum specific growth rate and doubling time for a
culture of bacteria from the diagram above.
Nutrient limitation Nutrient limitation
Nutrient limitation occurs when the growth of the organism is restricted by
a single nutrient, for example, C, N, P.
The way in which the specific growth rate falls from its maximum value
during the deceleration phase as the substrate concentrate becomes
limiting, was first reported by Jacques Monod (1910-1976) and is known
as the Monod equation:
s K
s
s
m
+
=
s Nutrient concentration [mg L
-1
]

m
Maximum specific growth rate [hr
-1
]
K
s
Saturation coefficient for the growth limiting nutrient, a
measure of the ability of the organism to uptake the nutrient
Nutrient limitation Nutrient limitation
Exercise: Calculate the maximum specific growth rate,
m
, if the glucose
substrate concentration is 100 mg C L
-1
. Assume K
s
is 5 mg C L
-1
, and is
0.2 h
-1
.
Production of Biomass in Production of Biomass in
Continuous Cultures Continuous Cultures
Our analysis of bacterial growth in fixed volume batches is only of limited
value in many engineering application, where continuous flow processes
are used (e.g. wastewater treatment)
In a cont. process biomass is retained inside a reactor and the stream to
be treated (e.g. wastewater, industrial effluent, contaminated groundwater)
is fed continuously to the reactor, and treated product is withdrawn at the
same rate.
Mass Balance on Biomass Mass Balance on Biomass
Although fresh nutrients are added continually, all other factors will remain
constant. We can define the mean residence time (or hydraulic retention
time, HRT) as:
Q
V
HRT =
V reactor volume [m
3
]
Q Feed flow rate [m
3
h
-1
]
Sometimes it is convenient to refer HRT to dilution rate, D, which is the
reciprocal of the HRT with units [h
-1
].
Mass Balance on Biomass Mass Balance on Biomass
Y Yield coefficient, kg biomass per kg nutrient
If D
m
, biomass will be retained in reactor
If D >
m
, biomass wash-out will occur
The mass of cells, m, produced in the reactor per unit time is given by:
( ) s s YQ m
i
=

Y Yield coefficient, kg biomass per kg nutrient

Q Feed flow rate [m
3
h
-1
]
s
i
Concentration of limiting nutrient in influent [kg m
-3
]
s Concentration of limiting nutrient in reactor [kg m
-3
]
Oxygen limitation Oxygen limitation
In order to achieve a high productivity from an aerobic continuous culture,
like the activated sludge process, it is necessary to run at a high dilution
rate, D (or low HRT), and high biomass production, m. In order to increase
m, a high concentration of the limiting nutrient, s
i
, is required. However, as
m is increased there will be a maximum value at which oxygen supply will
become limiting!
For aerobic organisms, a specific oxygen uptake rate (q
0
) can be
determined with the units of kg O
2
per kg cell and hr. The rate at which determined with the units of kg O
2
per kg cell and hr. The rate at which
oxygen is transferred into the water must exceed this minimum.
The equation for oxygen transfer into solution is given by:
n = k
L
a(c
DO
*-c
DO
)
n Rate of O
2
mass transfer [kg h
-1
]
k
L
a = absorption coefficient [m
3
h
-1
]
c
DO
Dissolved oxygen concentration in influent [mg L
-1
]
c
DO
* - Equilibrium DO concentration [mg L
-1
]
Oxygen limitation (contd) Oxygen limitation (contd)
A maximum cell concentration, m
max
/ V, can be determined for a given
absorption coefficient and permissible minimum dissolved oxygen (DO)
concentration, c
DO,min
:
( )
HRT q
c c a k
V
m
DO DO L
0
min ,
*
max

=
HRT q V
0
Exercise: Calculate the HRT at which a continuous flow biomass unit
must operate in order to achieve a maximum cell production of 28 kg
cells m
-3
h
-1
, within the limit of oxygen supply.
Given:
q
0
= 8 x 10
-2
kg O
2
(kg cells)
-1
h
-1
k
L
a = 250 h
-1
c
DO
* = 6 x 10
-3
kg m
-3
c
DO,min
= 1 x 10
-3
kg m
-3