1

CE 333: ENVIRONMENTAL ENGINEERING II

Lecture- 4: WASTEWATER TREATMENT (Part-2):
Biological (Secondary) Treatment Systems

Introduction:
The purpose of primary treatment is to remove suspended solids and floating material. In many
situations in some countries, primary treatment with the resulting removal of approximately 40 to 60%
of the suspended solids and 25 to 35%of BOD, together with removal of material from the wastewater,
is adequate to meet the requirement of the receiving water body.
If primary treatment is not sufficient to meet the regulatory effluent standards, secondary treatment
using a biological process is mostly used for further treatment due to its greater removal efficiency and
less cost than chemical coagulation.

Objectives of Secondary Treatment of waste water

Main objective
Secondary treatment processes are intended to remove most of the fine suspended and dissolved
degradable organic matter that remains after primary treatment (removal efficiency > 90%), so that the
effluent may be rendered suitable for discharge. Conventional secondary treatment can reduce the
BOD5 < 20mg/L and TSS < 30mg/L which is acceptable in most cases .

Second objective
The second objective of secondary treatment is the reduction of ammonia toxicity and nitrification
oxygen demand in the stream. This is achieved by oxidation of most of the ammonia to nitrate during
treatment (nitrification). This utilizes modified conventional biological treatment process.

Biological treatment processes provide similar biological activities to waste assimilation, which would
take place in the receiving waters, but in a reasonably shorter time.

Fundamentals of Biological Treatment

The basic mechanism of biological treatment process is the stabilization of wastewater biologically
using a variety of microorganisms. The wastewater is introduced into a specially designed bioreactor
where under aerobic or anaerobic conditions and organic matter is utilized by microorganisms such as
bacteria (aerobically or anaerobically), algae, and fungi (aerobically) etc. under favorable
environmental conditions, such as dissolved oxygen, nutrient, etc.

The microorganisms are able to decompose the organic matter through two different biological
processes (metabolism process):

i. Biological Oxidation (Catabolism)

This is the process of obtaining energy. The biological oxidation forms some end-products, such as
minerals, that remain in the solution and are discharged with the effluent.

COHNS (Organic matter) + O2 → CO2 + NH3 + Energy  + Other  end  products + Energy

bacteria

ii. Biosynthesis (Anabolism)

The biosynthesis transforms the colloidal and dissolved organic matter into new cells that form in turn
the dense biomass that can be then removed by sedimentation

COHNS (Organic matter) + O2 → C5H7NO2 (New  cells)

bacteria

Generally, the bacterial growth can be explained by the following simplified figure:

Organics + Bacteria + Nutrients + O2    → New Bacteria + CO2+ H2O + Residual Organics + Inorganics

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 2
Several bioenvironmental factors affect the activity of bacteria and the rate of biochemical reactions,
e.g, temperature, pH, F/M ratio etc. Therefore for efficient biological treatment design it is essential to
have detail information on the microorganisms involved in the treatment process.

Microbiology of Sewage
The microorganisms important for biological treatment of sewage and other biodegradable wastewater
are – bacteria, fungi, algae, protozoa, rotifers, crustaceans etc. Carbon is the basic building block for
microbial cell synthesis. A source of energy must be obtained from outside the cell to enable synthesis
to proceed. The relationship between the source of carbon and the source of energy for the
microorganism is important.

The goal in wastewater treatment is to convert both the carbon and the energy in the wastewater into
the cells of microorganisms, which can be removed from the water by settling or filtration. Therefore,
the processes are designed and operated to encourage the growth of organisms that use organic
material for both their carbon and energy source.

Classification of Microorganisms
(A) Classification by Energy and Carbon Source (metabolism)
These are further grouped into two categories- i. heterotrophs & ii. autotrophs

i) Heterotrophs –
A heterotroph is a group of organisms that obtain their food from other organisms and are not capable
of producing their own food. These organisms use organic material as a supply of both carbon (food)
and energy. Heterotrophs include all animals and fungi, some bacteria & many parasitic plants.

Some heterotrophs directly depend on autotrophs for their food like herbivores feeding on plants. Other
heterotrophs indirectly depend on the producers by feeding on the first type of heterotrophs. Most
heterotrophs depend on the process of photosynthesis in a number of different ways. In addition to
providing the energy and food, photosynthesis also provides oxygen tot eh heterotrophs. Heterotrophs
have greater importance in sewage treatment as they breakdown organic matter for their use.

Heterotrophs are further divided into two types based on the source of their energy- a)
photoheterotrophs which obtain their energy from light but depend on producers for their carbon source
and b) chemoheterotrophs, in turn, obtain both their energy and carbon from other producers.

Fig-1: Heterotrophs Fig-2: Autotrophs

ii) Autotrophs
An autotroph is a group of organisms capable of producing their own food by utilizing various
substances like water, sunlight, air, and other chemicals and use CO2 to supply their carbon needs.
The most commonly known autotrophs are plants; however, several other varieties of autotrophs are
found in nature, ranging from algae, phytoplankton, and some bacteria. Autotrophs are capable of
producing complex chemical structure from basic inorganic materials. Most autotrophs use
photosynthesis to convert solar energy to chemical energy, but various autotrophs also utilize other
processes like phototrophy and chemotrophy. Autotrophic organisms are of limited importance in
sewage treatment as they do not dependent on organic wastes for their metabolism.

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 3

(B) Classification by Oxygen Relationship

Microorganisms often also classified by their ability or inability to utilize oxygen in oxidation-reduction
reactions.

i) Obligate aerobes
Obligate aerobes are microorganisms that must have oxygen for their metabolism process. Most
eukaryotes are obligate aerobes and cannot survive without oxygen.

ii) Obligate anaerobes

Obligate anaerobes are microorganisms that cannot survive in the presence of oxygen. Wastewater
that is devoid of oxygen is called anaerobic.

iii) Facultative anaerobes

A facultative anaerobe is an organism which can survive in the presence of oxygen, can use oxygen in
aerobic respiration, but can also survive without oxygen via fermentation or anaerobic respiration.

iv) Aerotolerant anaerobes

Aerotolerant anaerobes do not use oxygen because they usually have a fermentative metabolism
(extraction of energy from carbohydrates in the absence of oxygen), but they are not harmed by the
presence of oxygen as obligate anaerobes are.

v) Microaerophiles
Microaerophiles are bacteria that require a minimum level of oxygen for growth, about 1%–10%, well
below the 21% found in the atmosphere.

Some Microbes of Interest in Wastewater Treatment

Bacteria
The highest population of microorganisms in a wastewater treatment plant will belong to the bacteria.
Bacteria are single celled prokaryotic microorganisms. They utilize soluble organic matter as food and
in general found whenever moisture and food source are available.
Their usual mode of reproduction is by binary fission. In this process a bacteria cell duplicates its
genetic material, or deoxyribonucleic acid (DNA), and then divides into two parts, with each new
organism receiving one copy of DNA.

Fig-3: Bacterial cell Fig-4: Binary fission

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 4
Bacteria are capable of storing food particles outside the cell by means of extracellular enzymes and
hence can remove soluble, colloidal and solid organic matter from wastewater. Bacteria are very
sensitive to light, air, water, food, temperature, pH and dissolved oxygen, etc. Among these, temp and
pH play a vital role in the life and death of bacteria. Most bacteria cannot tolerate pH levels above 9.5
or below 4.0.and optimum pH for growth lies between 6.5 and 7.5. The rate of reaction increases with
increase in temperature- doubling with about 100C rise in temperature depending on bacterial
characteristics.

Classification of Bacteria

1. Depending on Shape-
They have one of three basic shapes:
i. Bacillus (Rod shaped) : Width 0.5 to 1.0 μm, and length 1 to 3 μm
ii. Coccus (Spherical) : Diameter 0.5 to 1.0 μm
iii. Spirillum (Spiral) : Width 0.5 to 5 μm, and length 6 to 15 μm

2. Classification depending on Temperature

Temperature is the most important factor in biochemical reactions. Each species of bacteria
reproduces best within a limited range of temperatures. The rate of reaction increases with increase in
temperature - double with about 100C rise in temperature.
Four temperature ranges are used to classify bacteria:

Temperature (ºC)
Type
Range Optimum
Cryophilic
-2 to 30 12 to 18
Or Psychrophilic
Mesophilic 20 to 45 25 to 40
Thermophilic 45 to 75 55 to 65
60 to near
Hyperthermophiles 70 to 85
boiling

Bacteria will grow at slower rates over a larger range of temperatures and will survive at a very large
range of temperatures. For example, Escherichia coli, classified as a mesophile, will grow at
temperatures between 200C and 450 C and but will reproduce, although very slowly, at temperatures
down to 00C. If frozen rapidly, they and many other microorganisms can be stored for years with no
significant death rate. Once the optimum temperature range is exceeded, growth rate drops off rapidly
due to the denaturation of key proteins.

3. Classification by Oxygen Relationship

(definition discussed earlier)

Some important bacteria:

Pseudomonas: Reduce NO3 to N2, So it is very important in biological nitrate removal in treatment
works.
Zoogloea: Helps through its slime production in the formation of flocs in the aeration tanks.
Sphaerotilus natuns: Causes sludge bulking in the aeration tanks.
Bdellovibrio: destroy pathogens in biological treatment.
Acinetobacter: Store large amounts of phosphate under aerobic conditions and release it under an –
anaerobic condition so, they are useful in phosphate removal.
Nitrosomonas: transform NH4 into NO2-
Nitrobacter: transform NO2- to NO3-
Coliform bacteria: Coliform bacteria are found in soil, sewage and polluted water. They have their
origin in the intestinal tract of human beings and other warm blooded animals. The Coliforms are not
considered as pathogenic, but they are used as indicator of fecal contamination
CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 5
The most common type is E-Coli or Echerichia Coli, (indicator for the presence of pathogens). E-Coli is
measured in (No/100mL). Total Coliform (TC) and Fecal Coliform (FC) counts are made for water and
effluents to measure the bacteriological quality of water, wastewater and sewage

Examples of typical chemical reaction in some bacteria

Typical biochemical reactions representing aerobic and anaerobic metabolism of both autotrophic and
heterotrophic bacteria are presented below:

i. Heterotrophic aerobic bacteria:

C6H12O6 + 6O2 = 6 CO2 + 6H2O;

ii. Heterotrophic anaerobic bacteria:

C6H12O6 = 3 CH4 + 3CO2;
iii. Autotrophic- chemosynthetic-aerobic bacteria:
2NH4+ + 3O2 = 2NO2- + 4H+

iv. Autotrophic-chemosynthetic-anaerobic bacteria:

5S + 2H2O + 6NO3- = 5SO42- + 3N2 + 4H+

Metabolism of Bacteria
The sum total of all the chemical processes of the cell is called metabolism
It may be separated two components:

1. Catabolism for energy

it is the process of obtaining energy, and

2. Anabolism for synthesis

It is the process of synthesis of cellular components.

Both the catabolic processes and the anabolic processes are very complex and carried out through a
series of reactions catalyzed by enzymes. e.g. The general stages of catabolism of fats,
carbohydrates, and proteins Hydrolysis occurs in stage -1, which is the process of splitting of a polymer
by adding water to a covalent bond. The reaction is catalyzed by a hydrolyase enzyme.

3. Autolysis Process
When a microorganism cell has depleted its food reserves, it starts to oxidize itself, this process is
known as autolysis (=self-destruction), which leads to the death of bacteria.

Autolysis does not proceed to completion since approximately 20-25% of the cell mass is resistant to
aerobic degradation. From the metabolism process diagram shown below it is noted that one-third of
BOD is used in catabolic reactions and two-thirds in anabolic reactions.

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 6

Fig-6: Metabolism process diagram of bacteria

Fungi
Fungi are multi-cellular, non-photosynthetic, heterotrophic microorganisms. Most
fungi are strictly obligate aerobes that reproduce by a variety of methods including
fission, budding, and spore formation. They have ability to grow under low
moisture condition and can tolerate a relatively low pH range of 2 to 9, optimum is
5 to 6. Their cells require only half as much nitrogen as bacteria so that in a
nitrogen deficient wastewater, they predominate over the bacteria.

These qualities make fungi very important in biological treatment of some industrial waste, and
composting of solid waste. Being relatively large and filamentous shape, fungi tends to settle poorly
and undesirable in biological treatment process.

Algae
Algae are unicellular or multi-cellular, autotrophic photosynthetic microorganisms. Because of the
chlorophyll contained in most species, they produce oxygen through photosynthesis. In the presence of
sunlight, the photosynthetic production of oxygen is greater than the amount used in respiration. .

Photosynthesis:
CO2 + 2H2O CH2O + O2 + H2O

At night they use up oxygen in respiration

Respiration:
CH2O + O2 CO2 + H2O

If the daylight hours exceed the night hours by a reasonable amount, there is a net production of
oxygen.
Algae are of benefit in stabilization lagoons for wastewater treatment when they supply oxygen in
excess of respiration. Other than production of oxygen, they do not contribute to the stabilization of
waste because they use carbon dioxide or bicarbonates as a source of carbon rather than organic
carbon. Moreover, they are a liability when they leave in the lagoon effluent because they contribute to
the total suspended particulate concentration and may cause discharge limits to be exceeded.

Protozoa
Protozoa are motile, microscopic microorganisms that are usually single-celled.
The majority of protozoa are aerobic chemo-heterotrophs, although a few are
anaerobic. Protozoa are generally an order of magnitude larger than bacteria in
size, and often consume bacteria as an energy source. The protozoa act as
polishers of effluent from biological waste treatment processes by consuming
bacteria and particulate organic matter. They reproduce by binary fission.

Rotifers
Rotifers are aerobic, chemo-heterotrophic and multi-cellular organisms/animal.
These are multicellular organisms with well-developed tissues and organs. The
rotifer derives its name from the apparent rotating motion of two sets of cilia on
its head. The cilia provide mobility and a mechanism for catching food. For this
they are sometimes called "wheel animals".

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 7
Rotifers are very effective in consuming dispersed and flocculated bacteria, protozoa and small
particles of organic matter. Presence of rotifers indicates efficient aerobic purification process

Crustaceans
Crustaceans are heterotrophic and multi-cellular animal with hard
body or shells that includes shrimp, lobsters, and barnacles. .
They are normal occupants of natural waters and are a source of
food for fish and are not present in noticeable extent in biological
treatment of wastewater systems except in under-loaded lagoons.
Thus, but their presence is indicative of an effluent low in organic
matter and high in dissolved oxygen.

Viruses
Viruses are smallest biological unit containing all the information
necessary for their reproduction.
They are inter-cellular parasites and as such they require a host to
live in. Once they have a host, they redirect their complex
machinery to produce new viruses and eventually the host cell
ruptures releasing new viruses which go on infecting other cells.
Viruses are found in sewage and wastewater and are required to
be controlled by effective sewage treatment.

Pathogens
Pathogens are disease producing microorganisms . Some examples are given below:

Name of Pathogen Type of Organism Disease Caused

Bacillus Typhi Bacteria Typhoid fever
Salmonella Paratyphi Bacterial Paratyphoid fever
Endamoeba Hystolytica Protozoa Amoebic dysentery
Shigella Dysenteriea Bacteria Bacillary dysentery
Vibrio cholerae Bacteria Cholera
Gastroenteritis,
Enteric Viruses Virus
Jaundice

Favorable Environmental Factors for Bacterial Growth

Nutrients
Like most organisms, bacteria require nutrients to grow and maintain their cell functions. Bacteria
require relatively large amounts of carbon, nitrogen, hydrogen and oxygen; smaller amounts of
phosphorus, sulfur, potassium, calcium, iron and magnesium; and trace quantities of several other
elements (e.g. zinc, molybdenum). Most domestic wastewaters contain enough carbohydrates,
proteins, and fats, which supply most of the nutrients that bacteria require. It is generally industrial
wastewaters that lack sufficient nutrients.

Different bacteria have different nutritional requirement. With increase

in concentration nutrition, growth rate of bacteria increases up to
certain level and then growth rate remains constant irrespective of
nutrition addition.

The ideal formula for determining the nutritional needs of microorganisms, expressed in a ratio of
BOD:Nitrogen: Phosphorus (B:N:P) is 100:10:1. If any of these nutrients are lacking, operational
problems can result.

Temperature
Temperature affects the growth of bacteria by various ways. Below minimum temperature cell
membrane solidifies and become stiff to transport nutrients in to the cell, hence no growth occurs.
Whereas, above maximum temperature, cellular proteins and enzymes denatures, so the bacterial

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 8
growth ceases. Reactions are slower in colder temperatures and faster in warmer temperature.
Therefore more bacteria are required to do the same job during the winter

Fig-14: Effect of Temperature on microbial growth Fig-15: Effect of pH on microbial growth

pH
Most bacteria grow best around neutral pH values 6.5 - 7.0, but some thrive in very acid conditions
(alkaliphiles bacteria) and some can even tolerate a pH as low as 1.0 (acidophiles bacteria). Basically,
enzymes in bacteria are very much dependent on pH. Therefore, for effective microbial growth rapid
pH changes should be avoided.

Toxic elements
Presence of toxic elements (Chlorine compounds, Cr, Pb, Hg, Ag, etc) stops microbial growth
activities.

Dissolved Oxygen
This is the vital need for survival of aerobic microorganisms. Aerobic bacteria require 0.1 to 0.3 mg/L of
oxygen to function properly. Since the floc is made up of bacteria, a DO of around 2 mg/L should be
maintained in the bulk fluid surrounding the floc so that sufficient DO is available for bacteria in the
center of the floc. The floc solids will begin to break up and unfavorable bacteria that thrive under low
DO conditions may begin to develop.

Fig-17: Thioglycolate tubes test for determination of microbial oxygen requirements

The growth of bacteria with varying oxygen requirements in thioglycolate tubes is illustrated in
Figure 17. In tube A, all the growth is seen at the top of the tube. The bacteria are obligate (strict)
aerobes that cannot grow without an abundant supply of oxygen. Tube B looks like the opposite of
tube A. Bacteria grow at the bottom of tube B. Those are obligate anaerobes, which are killed by
oxygen. Tube C shows heavy growth at the top of the tube and growth throughout the tube, a typical
result with facultative anaerobes. The aerotolerant anaerobes in tube D are indifferent to the presence
of oxygen. Tube E on are microaerophiles bacteria that require a minimum level of oxygen for growth.

Ions and salt:

All bacteria requires metal ions such as K+, Ca ++, Mg++, Fe++, Zn++, Cu++, Mn++ etc to synthesize
enzymes and proteins.

Mixing
Mixing is required to bring the bacteria, oxygen and nutrients in contact with each other. Without
sufficient mixing, proper treatment will not take place.

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 9
The Bacterial Growth Pattern
The stabilization (decomposition) of sewage and industrial wastewater is related to the growth of
bacteria. The carbon and energy sources (wastes) in the wastewater for bacterial growth are referred
to as substrates. Bacteria utilize substrates and reproduce by binary fission process which may vary
from 20 minutes - days. But bacteria would not continue to divide indefinitely because of various
environmental limitations.

Generation time:
It is the time required for a bacterium to give rise to 2 daughter cells under optimum conditions. It is
also referred as population doubling time. Different types bacteria require different doubling time. e.g., -
—- Escherichia coli – 20 mins.
—- Mycobacterium tuberculosis – 20 hrs.
—- M.leprae – 20 days.

Estimation of Generation Time

The generation time can be calculated from the growth curve. The exactly doubled points from the
absorbance readings were taken and, the points were extrapolated to meet the respective time axis.

Generation Time
= (Time in minutes to obtain the absorbance 0.4) – (Time in minutes to obtain the absorbance 0.2)
= 90 – 60 mins
= 30 minutes

When bacteria come in contact with wastewater, they enter in to a new environment. Bacteria then
take some time to get acclimatized with new environment before starting binary fission. The general
growth pattern of bacteria in a fixed substrate environment has few distinct phases – lag phase, log
growth phase, declining growth, stationary and death phase. A general growth pattern for fission-
reproduction bacteria in a batch culture is sketched in figure below.

1. Lag Phase:
The initial stage of bacterial growth to get adjust with the new environment is termed as lag phase.
During lag phase of growth, bacteria remain active but do not reproduce. Necessary enzymes and
metabolites are built up to degrade substrates. In lag phase, cells are increased in size, but the
bacteria are not able to replicate and therefore no increase in cell mass.

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 10
The length of the lag phase depends directly on the previous growth condition of the organism. When
the microorganism growing in a rich medium is inoculated into nutritionally poor medium, the organism
will take more time to adapt with the new environment.

2. Log / Growth Phase:

As the bacterial get acclimated, they enter into progressive multiplication phase called the log phase.
Log phase of growth also is known as exponential or logarithmic phase. There are two significant
portions to the log phase of growth - i. substrate uptake and ii. synthesis of cells. During ―substrate
uptake,‖ bacterial cells simple become ―fat‖ due to the uptake (adsorption and absorption) of
substrates. The production of new daughter cells begins during ―synthesis of cells and rapid growth.‖
Generation time is constant for a given strain under same set of condition. Maximum rate of utilization
requires that other growth factors, e.g., oxygen, be supplied at a maximum rate and this may be
difficult to do.
Log growth phase coincides with the maximum rate of substrate (waste) removal, but not the optimum
zone of operation for waste treatment systems. Bacteria having a great deal of food & energy available
– become growing, motile, and extremely active. They got flagella, which enables them to swim to find
available food and hence are likely to be dispersed, and difficult to remove by sedimentation.

3. Declining Growth Phase:

As the level of food decreases, the growth rate slows as bacteria use up available substrates and living
space becomes limited. In this phase bacteria is approaching the carrying capacity or maximum
number of organisms that the biological treatment unit can support. Bacteria begin to conserve energy
and they lose the flagella. Since little food is available, they do not waste energy trying to swim to find
it. Next, they begin to form a thick outer slime layer.

4. Stationary Phase:
Growth ceases due to exhaustion of nutrients, accumulation of toxic metabolites and lowering of pH
and stationary growth or endogenous phase of growth is reached. This phase is indicates that the
bacterial population has reached the carrying capacity of the biological treatment unit.
At this stage bacteria start to produce secondary metabolites (a switch in energy and carbon flux away
from biomass production toward the production of small, bioactive molecules). Total cell count remains
same, i.e., the growth of new cells is offset by death of old cells and the number remains relatively
constant. As mobility ceases, bacteria bump into each other they stick together. At first they form small
clumps /flocs which eventually become large enough to settle.

5. Death Phase:
The depletion of nutrients and the subsequent accumulation of metabolic waste products and other
toxic materials in the media will facilitates the bacterium to move on to the Death phase. During this,
the bacterium completely loses its ability to reproduce. Individual bacteria begin to die due to the
unfavorable conditions.
The rate of death exceeds the rate of reproduction; cells also become old, die exponentially and lysis
(the disintegration of a cell by rupture of the cell wall or membrane) can occur in which the nutrients
from the dead cells diffuse out to furnish the remaining living cells as food.

Endogenous respiration
During both the stationary and death phases there are a substantial proportion of cells which neither
die nor subdivide. They exist by utilizing the intracellular food reserves laid down during exponential
growth; this process is known as endogenous respiration. In the endogenous growth period, the
microorganisms compete for the limiting substrate even to metabolize their own protoplasm.

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 11
Significance of bacterial growth analysis
The study of bacterial growth pattern is important as it helps to determine the hydraulic retention time
of wastewater in treatment basin which in turn depends on –

At which phase the rate of waste stabilization would be maximum??

In biological treatment, in addition to stabilization of biodegradable wastes newly created biomass also
need to be removed from the wastewater through sedimentation. Though substrate utilization is
maximum in exponential growth period, but as bacteria at this phase are highly active and motile –their
floc formation and sellting is very poor. During declining growth stage, activity ceases floc formation
increases which further grow in size during endogenous phase. Floc can settle out to the bottom of the
ASB, which leaves a clearer effluent with little BOD behind. Therefore a system should be run in the
―sweet spot‖ right between the declining growth phase and endogenous respiration phase for good
results.

Food : Microorganism Ratio (F:M)

Changes in F:M will cause of a change or upset in the biological activity in the secondary process and
may result in effluent quality problems. Actively growing bacteria consume at a rapid rate utilizing
oxygen at a rapid rate. Generally a higher uptake rate is associated with a higher F:M ratio and
younger sludge ages. A lower uptake rate is associated with a lower F:M ratio and older sludge ages.

The Table below shows overall waste stabilization and biomass settling efficiency at different growth
stage of bacteria:

Growth Waste Removal Bio-mass Settling

F/M ratio
Phase Rate Characteristics
Log Very High Maximum Dispersed, Poor
Declining
Moderate Good Good
Growth
Stationary Low Moderate Very good
Death Very low Negative Autolysis

Microbial Growth Kinetic Terminology

i. bCOD: Biodegradable COD
Since wastewater contains numerous substrates, the concentration of organic compounds is defined
by biodegradable COD. bCOD comprise soluble, colloidal, and particulate components.

ii. bsCOD: biodegradable soluble COD: Soluble components present in bCOD.

iii. TSS (total suspended solids) and VSS (volatile suspended solids):
It represents the biomass solids (insoluble) in the bioreactor.

iv. nbVSS: non-biodegradable volatile suspended solids

v. iTSS: inert inorganic total suspended solids

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 12
Biomass Yield
Biomass yield is defined as the ratio of the amount of biomass produced to the amount of substrate
(food) consumed

iomass produced gm
iomass yield,
ubstrate utili ed consumed gm

Since wastewater contains a large number of organic compounds, the yield is expressed in terms of
measurable parameters such as COD or BOD. Thus the yield would be:

iomass gm
iomass yield,
D or D gm

** For biodegradable part of COD it is expressed as bCOD or bsCOD

Estimating Biomass Yield and Oxygen Requirements

Cell Yield (for glucose used)
A stoichiometric relationship exists between the substrate removal, the amount of oxygen consumed,
and the observed biomass yield. Assuming organic matter can be represented as C 6H12 O6 (glucose),
the following equation can be written:

3C6H12O6 + 8O2 + 2NH3  2C5H7NO2 + 8O2 + 14H2O

3(180) 8(32) 2(17) 2(113)

The yield based on the glucose consumed cab be obtained as follows:

( ) ( )
--------- (1)
( ) ( )

Yield of bacterial cell = 0.42 gm cells/gm glucose used

Cell Yield (for COD based)

To express the yield in COD bases, the COD of glucose (i.e.,bCOD or bsCOD) must be determined:

C6H12O6 + 6O2 + 2NH3  6CO2 + 6H2O

(180) 6(32)

( ) ( )
( ) ( )

C --------- (2)

Now, from eq-1, Yield of bacterial cell

Therefore, COD based Theoretical Yield,

0.42 gm cells gm glucose

= 0.39 / 𝐂 𝐝
1.07 gm D gm glucose

Oxygen Requirement
Stoichiometric relation shows - 8 moles of oxygen are required for 3 moles of glucose

3C6H12O6 + 8O2 + 2NH3  2C5H7NO2 + 8O2 + 14H2O

3(180) 8(32) 2(17) 2(113)

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 13
Therefore,
Oxygen requirement for oxidizing per gm glucose-

2 used 8 32 gm 2 mol
0.47 gm
lucose 3 180 gm mol

Oxygen requirement for oxidizing per gm COD = 0.47 / 1.07 = 0.44 gm

Microbial Growth Kinetics

The relation between the specific microorganism growth rate (μ) and the substrate concentration (S) is
a valuable tool in biotechnology. This relationship is represented by a set of empirically derived
theoretical models. These models are nothing but mathematical expressions generated to describe the
behavior of a given system. The Monod model (1942) the most widely used empirical model to
describe microbial growth kinetics

μm
Specific growth rate μ ----- (3)
s
Where;
μ = specific microbial growth rate, gm VSS/gm VSS - d
μm = maximum microbial growth rate, gm VSS/gm VSS -d
Maximum biomass growth rate can be obtained from laboratory experiments using 2-fold dilution
(2FD) method or the relative rate to detection (RRD) method
S = growth limiting (biodegradable o.m.) substrate concentration, mg/L
X = biomass (microorganisms) concentration mg/L
Ks= Substrate saturation constant
= substrate concentration at one half the maximum substrate consumption rate (rsu), mg/L

Yield Co-efficient

Yield coefficient is the ratio of maximum mass of cells formed to the mass of substrate utilized, mg/mg
Monod also related the yield coefficient (Y) to the specific rate of biomass growth (μ) and the substrate
utilization rate (rsu)

rsu = -

rsu = rate of substrate consumption/ utilization. mg/L- d

= Total substrate utilized within total retention time

And, cell growth rate for biomass X,

rg = μ X = (mg/L-d) ----- (4)

rg = - Y rsu ----- (5)

Rate of soluble substrate consumption/utilization,

μm k
rsu - - ----- (6)
s s

Where;
k = max. specific substrate utilization rate, mg substrate/mg biomass – d

=
(μm = maximum microbial growth rate, gm VSS/gm VSS –d)
Ks= substrate concentration at one half the maximum growth rate , mg/L

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET
 14
Net growth rate
The relationship between cell growth rate and substrate utilization rate is given by: (not all subs. is
converted to cells)
rg = - Y rsu
But bacteria experience loss in growth rate due to decay and predation, this is termed as endogenous
decay. The death rate rd of cells is proportional to the cell concentration X, because cells die in
proportion to their number.
Thus,
rd = k d X -----(7)

Here, kd = endogenous decay coefficient, per day

Therefore, net biomass cell growth rate = growth rate - decay rate (mg cells/L-d)

rg’ = rg - rd

rg’ -Yrsu - kd X -----(8)

or, = - kd X -----(9)

CE-333: Lecture-4: Wastewater Treatment (Part-2) /Prepared by: Snigdha Afsana, Assist Prof, Dept. of Civil Engineering, BUET