LABORATORY MANUAL

ENVIRONMENTAL ENGINEERING LAB [CEL 411]

VII SEMESTER CIVIL ENGINEERING

DEPARTMENT OF CIVIL ENGINEERING

 Environmental Engineering Lab

LABORATORY MANUAL
ENVIRONMENTAL ENGINEERING LAB
[CEL 411]

VII SEMESTER CIVIL ENGINEERING

Name: -------------------------------------------------------------

Class: --------------------------------------------------------------

Reg. No: ----------------------------------------------------------

Year: ---------------------------------------------------------------

DEPARTMENT OF CIVIL ENGINEERING

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INSTRUCTIONS TO BE FOLLOWED BY STUDENTS

1. Dress code- Navy blue Shirt, black pants and Shoe. Boys should insert their
shirt and girls should put up their hair.
2. Bring with them their own lab manual, calculator and other accessories for
doing the experiment.
3. Study the manual thoroughly before entering the lab.
4. The squad leader should submit the list of instruments required with the
counter signature of the staff in charge of the class. They should check the
instruments before proceeding the practical class and report the damages if
any to the concerned laboratory staff. The squad will be responsible for any
loss or damage caused to any instrument after issue.
5. Handle the instruments with maximum care and if any defect or damage
happens to the instruments, the matter should be immediately reported to
the staff member in charge and they have to obey the directions of the staff
member.
6. Return the instruments 15 minutes before the expiry of the period
prescribed for the practical class.
7. The observations should be entered in the lab manual in ink in the class
itself; where corrections are made, the new reading is to be entered only
under the initials of the staff member. Once the experiment is over, the
observations are to be got counter signed by the staff member.
8. If any calculation has to be done, it should be finished during the practical
class itself, unless the staff member permits to complete it at a later stage.
9. Get the lab manual signed by the staff member before writing the record.
10.The record should be submitted within the prescribed time according to the
directions of the staff member in charge of the class.
11.Lab should be kept clean.
12. Do not operate the machine/ equipments in the absence of the laboratory
staff.

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COURSE OUTCOMES

Continuous Internal Evaluation Pattern:

Attendance: 15 marks
Continuous Assessment: 30 marks
Internal Test (Immediately before the second series test): 30 marks

End Semester Examination Pattern:

The following guidelines should be followed regarding award of marks

(a) Preliminary work: 15 Marks

(b) Implementing the work/Conducting the experiment: 10 Marks
(c) Performance, result and inference (usage of equipment and troubleshooting): 25 Marks
(d) Viva voce: 20 marks
(e) Record: 5 Marks

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SYLLABUS

1. Determination of pH, Electrical Conductivity and Turbidity*

2. Determination of TS, TDS and TSS, TVS *
3. Determination of Alkalinity and Acidity *
4. Determination of Hardness *
5. Determination of Chlorides
6. Determination of Total Iron
7. Determination of Biochemical Oxygen Demand*
8. Determination of Chemical Oxygen Demand*
9. Optimum Coagulant dosage*
10. Break point Chlorination *
11. Determination of Available Chlorine in a sample of bleaching powder
12. Determination of Sulphates
13. Determination of Fluoride
14. Determination of Dissolved Oxygen*
15. Determination of nitrates
16. Determination of phosphates
17. Determination of any two Heavy Metal concentration
18. Total coliforms *
Note: * mandatory

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INDEX

Sl. Experiments Page No.

No.
1 Determination of pH, Electrical Conductivity and Turbidity 7

2 Determination of TS, TDS and TSS 13

3 Determination of Alkalinity and Acidity 18

4 Determination of Hardness 21
5 Determination of Chlorides 24
6 Determination of Total Iron 26
7 Determination of Biochemical Oxygen Demand 28
8 Determination of Chemical Oxygen Demand 31
9 Optimum Coagulant dosage 33
10 Break point Chlorination 35

11 Determination of Available Chlorine in a sample of bleaching 38

powder
12 Determination of Sulphates 41

13 Determination of Fluoride 43

14 Determination of Dissolved Oxygen 45

15 Determination of nitrates 48

16 Determination of phosphates 50

17 Determination of any Heavy Metal (ARSENIC) concentration 52

18 Determination of total and fecal coliform in water 55

REFERENCES 58

Prepared by: Approved by:

Neethu Paul Radha Unnikrishnan M

Assistant Professor HOD
Department of Civil Engineering Department of Civil Engineering

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DETERMINATION OF pH
Exp. No.: 1.1

AIM:
To determine pH of a given water sample.
THEORY:

The term pH refers to measure of hydrogen ion concentration in a solution and defined as the
negative log of H+ ions concentration in water and wastewater. Values of pH, 0 to a little less
than 7 are termed as acidic and values of pH a little above 7 to 14 are termed as basic. When
the concentration of H+ and OH ions are equal then it is termed as neutral pH.
APPARATUS:
pH meter, Beaker
REAGENT:
Buffer solutions of known pH value
PROCEDURE:
Three major steps are involved in the experiment.
1. Preparation of Reagents
2. Calibrating the Instrument
3. Testing of Sample
STEPS:
• Perform calibration of pH meter using standard pH solutions. The calibration procedure
would depend on pHrange of interest.
• In a clean dry 100 ml beaker take the water sample and place it in a magnetic stirrer, insert
the Teflon coatedstirring bar and stir well.
• Now place the electrode in the beaker containing water sample and check for the reading in
pH meter. Wait untilyou get a stable reading.
• Take the electrode from the water sample, wash it with distilled water and then wipe gently
with soft tissue.

PRECAUTIONS:
1. Make sure to store a pH electrode in its wetting cap containing electrode fill solution (3 M
KCl, purchased or prepared by dissolving 22.37 g KCl into 100 mL DI water). Do not store
electrode in DI water.
2. Never touch the membrane of the glass electrode with anything else except soft tissue
paper since it is fragile and is easily ruined if scratched or bumped.
3. Always bring the standards and the sample to the same temperature before measurement
(preferably room temperature).

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RESULT:
pH of given water sample is……………….

DISCUSSION:
Determination of pH is one of the important objectives in biological treatment of the
wastewater. In anaerobic treatment, if the pH goes below 5 due to excess accumulation of
acids, the process is severely affected. Shifting of pH beyond 5 to 10 upsets the aerobic
treatment of wastewater. In these circumstances, the pH is generally adjusted by addition of
suitable acid or alkali to optimize the treatment of the wastewater. pH value or range is of
immense importance for any chemical reaction. A chemical shall be highly effective at a
particular pH. Chemical coagulation, disinfection, water softening and corrosion control are
governed by pH adjustment. Lower value of pH below 4 will produce sour taste and higher
value above 8.5 a bitter taste. Higher values of pH hasten the scale formation in water heating
apparatus and also reduce the germicidal potential of chlorine. High pH induces the formation
of tri- halomethanes, which are causing cancer in human beings.

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MEASUREMENT OF CONDUCTIVITY

Exp. No.: 1.2

AIM:
To calibrate conductivity meter and measure conductivity of given water sample.
THEORY:

Conductivity, k, is a measure of the ability of an aqueous solution to carry an electric current.

This ability depends on the presence of ions; on their total concentration, mobility, valence
and on the temperature of measurement. Since the charge on ions in solution facilitates the
conductance of electric current, the conductivity of a solution is proportional to its ion
concentration. Conductivity cells of different cell constants are chosen mainly to ensure that
the actual resistance between the plates, when the cell is immersed in the solution, is within
practical measurable range. It is customarily reported in micromhos per centimeter
(μmho/cm). Freshly distilled water has a conductivity of 0.5-2 μmho/cm.

APPARATUS:
1. Conductivity meter,
2. Glassware (Beaker)
REAGENT:

 Conductivity water: Pass distilled water through a mixed bed deionizer and discard
first 1000 mL of water. Conductivity of this water should be less than 1 μmho/cm.
 Standard potassium chloride solution, KCl, 0.0100M: Dissolve 745.6 mg anhydrous
KCl in conductivity water and dilute to 1000 mL at 25°C. This is the standard
reference solution, which at 25°C has a conductivity of 1412 μmhos/cm.
PROCEDURE:

Calibration and measurement of sample:

1. Rinse the electrode thoroughly and then blot and dry.
2. Immerse the electrode in 0.01M KCl solution and the conductivity should be 1413 micro
mhos at 25°C Temperature. Adjust the knob to get the accurate value.
3. Now the instrument is calibrated.
4. Remove the cell from KCl solution and wash it thoroughly with distilled water.
5. Immerse the electrode/cell in the unknown sample whose conductivity is to be determined.
6. Note down the instrument reading.

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OBSERVATION TABLE:

SAMPLE NO CONDUCTIVITY TEMPERATURE

RESULT:
1. The conductivity of water sample A is ________
2. The conductivity of water sample B is ________
3. The conductivity of water sample C is ________

DISCUSSION:

Conductivity can tell us a lot about water quality. Drastic changes in conductivity can be used
as an indicator of poor water quality from runoff or other sources of pollution. As salts and
other inorganic chemicals dissolve in water, they break down into ions. When ions and
dissolved salts are present in the water, they increase the ability to conduct electricity,
therefore as salinity increases in water, so does the electrical conductivity (EC). Pure
(distilled) water is a good example of a poor conductor because it does not contain any
dissolved salts that provide ions to conduct electricity. Water contains certain elements or
chemicals that affect water quality, these include sodium, magnesium, calcium, and chloride.
As salinity and temperature affect conductivity, it is also important to measure these
parameters; as salinity and temperature increase, conductivity also increases. When an
increase or decrease is detected on a conductivity meter, it can indicate pollutants, affecting
the quality of the water. When a sewage leak or agricultural runoff occurs, it increases the EC
because of additional ions such as chloride, phosphate, and nitrate, whereas, events such as an
oil spill decrease the EC, however, both have a negative impact on water quality.

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DETERMINATION OF TURBIDITY OF WATER

Exp. No.: 1.3

AIM:
To determine the turbidity of given water sample.
THEORY:
Turbidity is the technical term referring to the cloudiness of a solution and it is a qualitative
characteristic which is imparted by solid particles obstructing the transmittance of light
through a water sample. Turbidity often indicates the presence of dispersed and suspended
solids like clay, organic matter, silt, algae and other microorganisms. So in short turbidity is
an expression of the optical property that causes light to be scattered and absorbed rather than
transmitted in straight lines through the sample. Turbidity is based on the comparison of the
intensity of light scattered by the sample under defined conditions with the intensity of the
light scattered by a standard reference suspension under the same conditions. The turbidity of
the sample is thus measured from the amount of light scattered by the sample taking a
reference with standard turbidity suspension. The higher the intensity of scattered light the
higher is the turbidity. Formazin polymer is used as the primary standard reference
suspension.
APPARATUS: Turbidity Meter

REAGENT: Formazin Polymer standards

SAMPLE HANDLING AND PRESERVATION:

Water samples should be collected in plastic cans or glass bottles. All bottles must be cleaned
thoroughly and should be rinsed with turbidity free water. Volume collected should be
sufficient to ensure a representative sample, allow for replicate analysis (if required), and
minimize waste disposal. No chemical preservation is required. Keep the samples at 4°C. Do
not allow samples to freeze. Analysis should begin as soon as possible after the collection. If
storage is required, samples maintained at 4°C may be held for up to 48 hours.

PRECAUTIONS:
The following precautions should be observed while performing the experiment:
 The presence of coloured solutes causes measured turbidity values to be low. Precipitation
of dissolved constituents (for example, Fe) causes measured turbidity values to be high.
 Light absorbing materials such as activated carbon in significant concentrations can cause
low readings.
 The presence of floating debris and coarse sediments which settle out rapidly will give low
readings. Finely divided air bubbles can cause high readings.

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PROCEDURE:
1. For testing the given water sample first the reagents are to be prepared. turbidity meter is
required to be calibrated. Then the
2. To the sample cells, add sample water up to the horizontal mark, wipe gently with soft
tissue and place it in the turbidity meter. Cover the sample cell with the light shield.
3. Check for the reading in the turbidity meter. Wait until you get a stable reading.

OBSERVATION TABLE:

RESULT:
1. The turbidity of water sample A is ________
2. The turbidity of water sample B is ________
The turbidity of water sample C is ________

DISCUSSION:

According to WHO standard 5 NTU (Nephelometric turbidity unit) is suggested as the

turbidity limit for drinking water, while 1 NTU is recommended to achieve adequate
disinfecting safety. According to Bangladesh Environment Conservation Rules (2023),
drinking Water standard for Turbidity is 5 NTU.

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TOTAL SOLIDS (TS)

Exp. No.: 2.1

AIM:
To determine total solids of given sample.
THEORY:
Total solids are determined by evaporating a well-mixed sample and dried to constant weight
in an oven at 103 to 105°C and weighing the dry residue left. The increase in final weight
than the initial weight of crucible indicates total solids.

APPARATUS:

Crucible, Muffle furnace, measuring cylinder, weight balance, oven, desiccators.

PROCEDURE:

1. Take the weight of empty crucible

2. Take 50 ml of given water sample in a crucible.
3. Evaporate the sample at 103 to 105°C for 24 hrs.
4. Again ignite the crucible for 15-20 minutes in a muffle furnace whose temperature is
maintained to 550°Cuntil the constant weight is attained. Cool it.
5. Note down the weight of crucible with residue.
OBSERVATION:
1. Weight of empty crucible (B):…………………. gm
2. Weight of empty crucible with residue (A):……………… gm.
CALCULATION:
Total solids (TS) mg/l = {(A-B) x 1000 x 1000} / ml of sample (V),
where,
A: Final weight of the crucible with residue in gm
B: Initial weight of the empty crucible in gm.
V: Volume of sample taken in ml.
RESULT:-
The amount of total solids determined from the given water sample is …….mg/l.

DISCUSSION:

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Total solids determination is used to assess the suitability of potential supply of water for
various uses.
TOTAL SUSPENDED SOLIDS (TSS) AND TOTAL DISSOLVED
SOLIDS (TDS)

Exp. No.: 2.2

AIM:
To determine total suspended solids from given sample.
THEORY:
The residue which remains on filter paper after filtration. The dry weight of this residue is
termed as total suspended solids.
APPARATUS:

Whatman filter paper no. 44, measuring cylinder, weight balance, oven, desiccator, funnel,
tripod stand.
PROCEDURE:

1. Take a tripod stand and put it on the table to which funnel is placed.
2. Take initial weight of Whatman filter paper as (W1)
3. Now place the Whatman filter paper three folded on the funnel.
4. Pour the well mixed sample on the Whatman filter paper.
5. Wait for some time so as to drain out the water sample completely.
6. Now remove the Whatman filter paper and keep it in the oven. Care should be taken to see
that the filterpaper should not be scratched.
7. Now take the final weight of the filter paper along with residue as (W2).
OBSERVATION:
1. Initial weight of Whatman filter paper (W1): ……………………gm.
2. Final weight of Whatman filter paper with residue (W2): …………..gm.
CALCULATION:
Total suspended solids (TSS) mg/l = { (W2-W1) x 1000 x 1000 } / ml of sample (V)
where,
W1: Initial weight of filter paper in gm.
W2: Final weight of filter paper with residue in gm
V = Volume of Sample taken
RESULT:-
The amount of total suspended solids determined from the given water sample is……… mg/l

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DISCUSSIONS:
1. Suspended solid material may be objectionable in water. Organic suspended are degraded
anaerobically mayrelease obnoxious odors.
2. Measures the quality of the wastewater influent and effluent.
3. Extremely valuable in the analysis of polluted waters.

APPLICATION OF DATA:
1. To evaluate the strength of domestic wastewater.
2. To determine the efficiency of treatment units.

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TOTAL DISSOLVED SOLIDS (TDS)

Exp. No.: 2.3

AIM:
To determine total dissolved solids from given sample.
THEORY:
The residue which remains on filter paper after filtration. The dry weight of this residue is
termed as total suspended solids.
APPARATUS:

Whatman filter paper no. 44, measuring cylinder, weight balance, oven, desiccator, funnel,
tripod stand.
PROCEDURE:

1. Take a tripod stand and put it on the table to which funnel is placed.
2. Take initial weight of Whatman filter paper as (W1)
3. Now place the Whatman filter paper three folded on the funnel.
4. Pour the well mixed sample on the Whatman filter paper.
5. Wait for some time so as to drain out the water sample completely.
6. Now remove the Whatman filter paper and keep it in the oven. Care should be taken to see
that the filterpaper should not be scratched.
7. Now take the final weight of the filter paper along with residue as (W2).
OBSERVATION:
1. Initial weight of Whatman filter paper (W1): ……………………gm.
2. Final weight of Whatman filter paper with residue (W2): …………..gm.
CALCULATION:
Total suspended solids (TSS) mg/l = { (W2-W1) x 1000 x 1000 } / ml of sample (V)
where,
W1: Initial weight of filter paper in gm.
W2: Final weight of filter paper with residue in gm
V = Volume of Sample taken
TDS (mg/l) = TS – TSS

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RESULT:-
The amount of total dissolved solids determined from the given water sample is……… mg/l

DISCUSSIONS:
1. TDS stands for total dissolved solids, and represents the total concentration of dissolved
substances in water.
2. An elevated total dissolved solids (TDS) concentration is not a health hazard. The TDS
concentration is a secondary drinking water standard and, therefore, is regulated because it is
more of an aesthetic rather than a health hazard.
3. High concentration produces hard water, which leaves deposits and films on fixtures and
on the insides of hot water pipes and boilers

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DETERMINATION OF ALKALINITIY & ACIDITY

Exp. No.: 3

AIM:
To determine alkalinity of a given water sample
THEORY:
Alkalinity is primarily a way of measuring the acid neutralizing capacity of the water. In
other words, its ability to maintain a relatively constant pH. The ability of natural water to act
as a buffer is controlled in part by the amount of carbonate ion in solution. Carbonate ion and
calcium ion both come from calcium carbonate or limestone. So water that comes in contact
with limestone will contain high levels of both Ca++ and CO - and have elevated hardness
and alkalinity.
The alkalinity of water is a measure of how much acid it can neutralize. When a water sample
that has a pH of greater than 4.5 is titrated with acid to a pH 4.5 end point, all OH- , CO3 2-
and HCO3 - will be neutralized. For pH more than 8.3, add phenolphthalein indicator, the
colour changes to pink colour. This pink colour is due to presence of hydroxyl ions. If
sulphuric acid is added to it, the pink colour disappears i.e. OHions are neutralized. Then add
mixed indicator, the blue colour appears. While adding acid, the colour changes to orange,
this colour change indicates thatall the CO3 2- & HCO3 - has been neutralized. This is the
end point.
APPARATUS:

Burette, pipette, Conical flask, Beakers, Burette stand and Clamp

CHEMICALS:
Sodium carbonate, Standard Sulphuric acid, Phenolphthalein and Methyl orange indicator,
Methyl red & Bromocresol green
PROCEDURE:
The procedure involves two phases.
1. Preparation of reagents
2. Testing of water sample
PREPARTION OF REAGENTS:
The following reagents are required to be prepared.
• Sulphuric acid solution (0.02 N)
• Phenolpthalein Indicator
• Mixed Indicator

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Sulphuric acid solution (0.02 N)

Take 500 ml of distilled water in a 1000 ml standard flask. Exactly measure 28 ml of
concentrated sulphuric acid and add slowly along the sides of the standard flask. Then make
up the volume up to the mark. The strength of this solution is 1 N. For preparing 0.02 N
sulphuric acid solution, exactly measure 20 ml of 1 N solution and make up to1000 ml in a
standard flask.
Phenolphthalein Indicator
For preparing 0.02 N sulphuric acid solutions, exactly measure 20 ml of 1N solution and
make up to 1000 ml in astandard flask. Weight 1 gm of phenolphthalein and add 100 ml of
95% ethyl alcohol or 100 ml of distilled water.
Mixed Indicator
Dissolve 100 mg bromocresol green and 20 mg methyl red in 100 ml 95% ethyl alcohol or
100 ml of distilled water.

TESTING OF WATER SAMPLE:

Using a measuring cylinder exactly measure 100 ml of sample and pour it in a 250 ml of
conical flask. Fill the burettewith 0.02N sulphuric acid. Add few drops of phenolphthalein
indicator to the conical flask. If the contents in the conical flask turns pink then titrate it
against 0.02N sulphuric acid till the pink colour disappears. Note down the titrate value (V1).
The value of titration is 0.5 ml. This value is used in calculating the phenolphthalein
alkalinity. To the same solution in the conical flask add few drops of mixed indicator and the
solution turns blue. Continue the titration from the point where stopped. Titrate till the
solution become red. The entire volume (V2) of sulphuric acid noted down and is
accountable in calculating the total alkalinity. Repeat the titration from concordant values.
Burette solution: Sulphuric acid solution
Pipette solution: Sample
Indicator: Phenolphthalein Indicator, Mixed Indicator
OBSERVATION AND CALCULATION:
Calculation of Phenolphthalein Alkalinity
Burette solution: Sulphuric acid solution
Pipette solution: Sample
Indicator: Phenolphthalein Indicator
End point: Disappearance of pink colour

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Calculation of Total Alkalinity

Burette solution: Sulphuric acid solution
Pipette solution: Sample
Indicator: Mixed Indicator
End point: Appearance of red colour

RESULT:
Total Alkalinity present in water sample is………….
DISCUSSIONS:

Large amount of Alkalinity imparts bitter taste in water. The principal objection of alkaline
water is the reactions that can occur between alkalinity and certain cations in water. The
resultant precipitate can foul pipes and other accessoriesof water distribution systems.
a. Chemical coagulation of water and waste water – to neutralize acids produced during
flocculation, the sample should be alkaline as otherwise further floc formation slowly ceases.
b. Corrosion control: to control the corrosion due to acids, natural water is rendered to
alkaline.
c. Effluents of wastewater: wastewater containing excess caustic (hydroxide) alkalinity is not
to be discharged into natural water bodies or sewers.

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DETERMINATION OF HARDNESS IN WATER

Exp. No.: 4

AIM:
To determine hardness of given water sample
THEORY:
Hardness is caused by divalent metallic cations. The principal hardness causing cations are
calcium, magnesium, strontium, ferrous and manganese ions. The major anions associated
with these cations are sulphates, carbonates, bicarbonates, chlorides and nitrates. The
hardness of water varies considerably from place to place. In general, surface water is softer
than groundwater. The hardness of water reflects the nature of the geological formations with
which it has been in contact. The total hardness of water is defined as the sum of calcium and
magnesium concentrations, both expressed as calcium carbonate, in mg/l. Hardness are of
two types, temporary or carbonate hardness and permanent or non-carbonate hardness.
Temporary hardness is one in which bicarbonate and carbonate ion can be precipitated by
prolonged boiling. Non-carbonate ions cannot be precipitated or removed by boiling, hence
the term permanent hardness. IS value for drinking water is 300 mg/l as CaCO3.
EDTA (Ethylenediamine tetra acetic acid) forms colorless stable complexes with Ca2+ and
Mg2+ ions present in water at pH = 9-10. To maintain the pH of the solution at 9-10, buffer
solution (NH4Cl + NH4OH) is used. EriochromeBlack-T (E.B.T) is used as an indicator. The
sample of hard water must be treated with buffer solution and EBT indicator which forms
unstable, wine-red colored complexs with Ca2+ and Mg2+ present in water.
APPARATUS:

50 ml Burette, 20 ml Pipette, 250 ml Conical flask, 100 ml Beaker, 250 ml beaker, Glass
funnel.
REAGENTS:
EDTA solution, Standard CaCO3 solution, Eriochrome Black–T indicator, Buffer solution.
PROCEDURE:
1. Standardization of EDTA
(i) Pipette out 20 ml of standard hard water into a conical flask.
(ii) Add 5 ml of buffer solution and few drops of Eriochrome Black-T. The
indicator, which is originally bluecolor would acquire a wine-red color.
(iii) Titrate with EDTA solution taken in the burette, till the wine red color changes
to blue which is the endpoint. Let the burette reading of EDTA be V2 ml
2. Determination of Total hardness
Repeat the above titration method for sample hard water instead of standard hard
water. Let the burette reading of EDTA be V3 ml.
3. Determination of Permanent hardness
Take 100 ml of sample hard water in 250 ml beaker. Boil it to remove temporary

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hardness to about half of this volume and cool to room temperature. Filter through
filter paper to remove insoluble CaCO3 and MgCO3. Make up the volume to the
original 100 ml by adding distilled water. Now pipette out 20 ml of this solution into a
clean conical flask. Then repeat the process of titration steps as mentioned above. Let
the burette reading of EDTA be V4 ml.
OBSERVATIONS:

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RESULT:
Total Hardness present is a water sample is……………..
DISCUSSIONS:
Hard water is as satisfactory for human consumption as soft waters. Because of their action
with soap, however, their use for cleansing purpose is quite unsatisfactory, unless soap costs
are disregarded. Soap consumption by hard waters represents an economic loss to the water
user. Sodium soaps react with multivalent metallic cations to form a precipitate, thereby
losing their surfactant properties. In recent years these problems have been largely alleviated
by the developments of soaps and detergents that do not react with hardness. Boiler scale, the
result of the carbonate hardness precipitation, may cause considerable economic loss through
fouling of water heater and hot water pipes. Change in pH in the water distribution systems
may also result in deposits of precipitates. Bicarbonates begin to convert to the less soluble
carbonates at pH values above 9.0. Magnesium hardness, particularly associated with the
sulfate ion has a laxative effect on persons unaccustomed to it. Magnesium concentrations of
less than 50 mg/l are desirable in potable waters, although many public water supplies exceed
the amount. Calcium hardness presents no public health problem. In fact, hard water is
apparently beneficial to the human cardiovascular system.

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DETERMINATION OF CHLORIDE IN WATER

Exp. No.: 5

AIM:
To determine chloride of given water sample
THEORY:
Chlorides occur in all natural waters in widely varying concentration; the chloride content
normally increases as the mineral content increases. Chloride ion may be present in
combination with one or more of the cations of calcium, magnesium, iron and sodium.
Chlorides of these minerals are present in water because of their high solubility in water.
Each human being consumes about six to eight grams of sodium chloride per day, a part of
which is discharged through urine and night soil. Thus, excessive presence of chloride in
water indicates sewage pollution. IS value for drinking water is 250 to 1000 mg/l.
The Mohr Method uses silver nitrate for titration (normality: 0.0141) (method applicability:
0.15 to 10 mg/L chloride ions). This corresponds to 1 ml of 0.0141 equals to 1 mg chloride in
solution. The silver nitrate solution is standardized against standard chloride solution,
prepared from sodium chloride (NaCl). During the titration, chloride ion is precipitated as

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white silver chloride (Eq.1):

The indicator (potassium chromate) is added to visualize the endpoint, demonstrating

presence of excess silver ions. In the presence of excess silver ions, solubility product of
silver chromate exceeded and it forms a reddish-brown precipitate (Eq.2). This stage is taken
as evidence that all chloride ions have been consumed and only excess silver ions have
reacted with chromate ions:

APPARATUS: Burette, conical flask, pipette, measuring cylinder.

REAGENTS: Potassium chromate indicator solution, standard silver nitrate titrant.

PROCEDURE:
1. Take 25 ml sample in a conical flask. Measure sample pH.
2. Add 1.0ml potassium chromate indicator solution,
3. Titrate with standard silver nitrate solution to pinkish yellow end point and note down
volume of titrant used.Also measure sample pH.
4. Calculate chloride ion concentration using Eq. (3):

Where: A = volume of titrant used,

N is normality of silver nitrate (here we used N/71 or 0.0141 N),
V sample is volume of sample used (ml).
RESULT:
Chloride present in water sample is………………

DISCUSSIONS:
Chlorides in reasonable concentrations are not harmful to human. At concentrations above
250 mg/l they give a salty taste to water, which is objectionable to many people. For this
reason, chlorides are generally limited to 250 mg/ l in supplies intended for public use. In
many areas of the world where water supplies are scarce, source be containing as much as
2,000 mg/l are used for domestic purposes without the development of adverse effects, once
the human system becomes adapted to the water.

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DETERMINATION OF TOTAL IRON IN WATER

Exp. No.: 6

AIM:
To determine total iron content of given water sample
THEORY:
Iron may be present in two forms, namely the reduced form (ferrous, Fe2+) and the fully
oxidized form (ferric, Fe3+). Ferric iron is seldom found in true solution in natural waters,
unless they are highly acidic, because of the formation of insoluble ferric hydroxides. Ferrous
iron is more likely to be found in true solution, although it is easily oxidized to the ferric state
and precipitated in alkaline waters as ferric hydroxide. Since some iron may exist as iron
hydroxide precipitates, therefore it is necessary to bring precipitated form(s) of iron back in
'to solution before oxidizing total iron content in water. For this hydrochloric acid is added to
the test sample to dissolve the insoluble ferric forms. For determination of total iron by the
following procedure, it must be ensured that all iron exists in ferric form (Fe3+). This is most
readily accomplished by using potassium permanganate, an oxidizing agent.

Ferric iron is determined by producing a red-colored iron compound, ferric thiocyanate, by

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the addition of potassium thiocyanate.

The quantity of ferric iron is determined by comparison with the red color produced by
standard iron solutions.
APPARATUS:
1. Nessler tube
2. Measuring cylinder
3. Dropper
REAGENTS:
1. Hydrochloric acid
2. Potassium permanganate solution
3. Potassium thiocyanate solution
4. Standard iron solution
PROCEDURE:
1. Place 100 mL of the water sample in a Nessler.
2. Add 5 mL of dilute hydrochloric acid
3. Add two drops of potassium permanganate solution
4. Add 5 mL of potassium thiocyanate solution. The solution would turn brown if iron is
present.
5. Compare the brown color formed with the standard prepared as follows:
a. Add 100 mL of distilled water in a Nessler tube
b. Add 5 mL of the dilute hydrochloric acid.
c. Add two drops of potassium permanganate solution.
d. Add 5 mL of potassium thiocyanate solution.
e. Add 0.2 mL at a time of the standard iron solution until the color of the standard and the
sample match.
CALCULATION:
Total iron concentration of test sample (mg/L) =………..

OBSERVATIONS:

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RESULT:
Total iron concentration of test sample (mg/L) =………..

DISCUSSIONS:
According to Bangladesh Environment Conservation Rules (2023), drinking water standard
for iron is 0.3 - 1.0 mg/l.

DETERMINATION OF BIOCHEMICAL OXYGEN DEMAND (BOD)

Exp. No.: 7

AIM:
To determine bod of given sample
THEORY
The BOD is an empirical test which measures the oxygen required by the microorganisms for
the biochemical degradation of organic matter to carbon dioxide and water at 20°C
temperature. The test consists of determination of DO prior to following period of 5 days.
The difference between first day DO and fifth day DO is the amount of BOD.
APPARATUS:
BOD bottles (capacity 300 ml), BOD Incubator, burette, pipettes, conical flask, burette
stands, tile, measuring cylinder, weight balance, glass rod, beakers.
REAGENTS:

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PROCEDURE:
1. Take the sample in 2 BOD bottles.
2. Fill another two BOD bottles with distilled water.
3. Add 1 ml each of phosphate buffer, magnesium sulphate, calcium chloride, and ferric
chloride solutions in all above bottles.
4. Find immediately DO of the sample and distilled water on 1st day.
5. Incubate at 20°C for 5 days the other two bottles. Tightly stopper to prevent any air entry
into the bottles.
6. Determine DO content in the incubated bottles at the end of 5 days by using DO estimation
method.
OBSERVATION:
1. In burette: 0.025N sodium thiosulphate solution
2. In conical flask: 50 ml sample + indicator
3. Indicator: Starch
4. End Point: Blue to Colourless

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SOURCE:
1. Wastewater
2. Tap water
3. Drinking water
4. Distilled water
OBSERVATION TABLE:

Sl Source Burette reading Difference Average

No

CALCULATIONS:

RESULT:
The amount of BOD determined from the given sample is ________ mg/l.
DISCUSSIONS:

1. It is principle test which gives biodegradability of a sample and the strength of waste.
Hence the amount of pollution can be measured.
2. Efficiency of any treatment plant can be judged by considering influent BOD and effluent
BOD and so also the organic loading on the unit.
3. Data from BOD tests are used for the development of engineering criteria for the design of
wastewater treatment plants

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CHEMICAL OXYGEN DEMAND (COD)

Exp. No.: 8

AIM:
To determine COD of given sample
THEORY
The organic matter present in sample gets completely oxidized by K2Cr2O7 in presence of
H2SO4 to form CO2 andH2O. The K2Cr2O7 gives the O2 required for the oxidation of
organic matter.
APPARATUS:

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COD Digester, Spectrophotometer, Measuring cylinder, Beakers, Volumetric flask.

REAGENTS:

REACTION:

PROCEDURE:
1. Take COD vials with stopper.
2. Add 2 ml of sample in the COD vials.
3. Add 1.5 ml of potassium dichromate reagents.
4. Add 3.5 mal of sulphuric acid reagent.
5. COD the vials become hot.
6. Cap the vials tightly.
7. Switch on the COD digester and fix the temperature at 1500°C and set the time at 2 hours.
8. Now place the COD vials into a COD digester and heat for 2 hours.
9. The digester automatically switches off. Then remove the vials from the digester and take
readings on spectrophotometer.

RESULTS:
The amount of COD determined from the provided water sample is………… mg/l.
DISCUSSION:
1. COD values are practically important in the survey designed to determine and control the
losses to sewer systems.
2. The ratio of BOD to COD is useful to assess the amenability of water for biological
treatment.
3. It is useful to assess strength of water, which contain toxins and biologically resistant
organic substance.
4. For domestic and some industrial waste water, COD value is about 2.5 times BOD values.

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OPTIMUM DOSAGE OF COAGULANT

Exp. No.: 9

AIM:
To find the optimum dose of coagulant required for treating the given turbid water sample.
THEORY
Treatment of water by coagulation involves –
(1) Determination of optimum dose of coagulant by jar test.
(2) Determination of power input for the flocculator. In the class jar test to determine

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optimum coagulant dose will be carried out, it is important to determine the optimum dose to
avoid charge reversal and resuspension colloids. Optimum coagulant dose is considered as
the amount of coagulant which produces water with lowest turbidity.
APPARATUS:
1. Jar test Apparatus.
2. pH meter.
3. One-liter beakers - 6 Nos.
4. Graduated pipette.
5. Turbidity meter.
REAGENTS:
Alum solution- Dissolve 1 g of fresh alum in 1000 ml of distilled water.
PROCEDURE:
1. Determine pH and turbidity of the water to be treated. You may be instructed to determine
color and arsenic concentration of the water to be treated (if removal efficiencies of these
parameters are to be determined).
2. Fill six 1000 mL beakers each with 500 mL water to be treated,
3. Add required (as instructed by teacher) coagulant (standard alum solution) to each beaker.
4. Mix the samples in the beaker with the help of the stirring device. Subject the samples to
one minute of rapid mixing followed by 15 minutes of slow mixing (about 40 rpm).
5. Allow the flocs to settle down for about 15 minutes. Observe the characteristics of the flocs
and the settling rates.
6. Collect the supernatant from each beaker and measure pH and turbidity of each. You may
be instructed to measure color and arsenic concentration of the supernatant samples (if
removal efficiencies of these parameters are to be determined).
7. Plot pH versus alum dose in a graph paper and observe effect of alum dose on pH.
8. Plot turbidity (NTU) versus the coagulant (alum) dose (mg/L) in a graph paper. Determine
optimum dose of alum from this plot.

OBSERVATIONS AND CALCULATIONS:

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RESULT:
Ideal dosage of coagulant (mg/l) =………..

DISCUSSIONS:
Besides efficient removal of turbidity and color, coagulation with alum and ferric chloride or
ferric sulfate is also widely used for removal of heavy metal ions (e.g., lead, arsenic) from
water. In this process heavy metal ions are primarily removed by adsorption (and Subsequent
precipitation) onto coagulated flocs of metal (either aluminum or iron) hydroxides.
Coagulation with alum and ferric chloride / sulfate is being successfully used for removal of
arsenic from water.

BREAK POINT CHLORINATION

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Exp. No.: 10

AIM:
To determine the residual chlorine in a given water sample
THEORY
Break Point Chlorination
If chlorine is added to water containing reducing agents and ammonia (either naturally
present or added to water to produce combined chlorine), a hump-shaped breakpoint curve is
produced as shown in following figure. The different segment of the curve is described as
follows:
a. If the water is free of ammonia and other compounds that may react with chlorine, the
application of chlorine will yield free available chlorine residual of same concentration. This
is denoted by the ‘no demand line’ or the "zero demand line" (see Fig.).
b. Chlorine first reacts with reducing agents such as H2S, Fe-2+, Mn2+ and develops no
measurable residual as shown by the portion of the curve from Origin up to point A.

c. Addition of chlorine beyond point A results in forming mainly mono- and di- chloramines.
With mole ratios of chlorine to ammonia up to 1:1 [i.e., C12:NH3-N = 1:1], both mono and
di-chloramines are formed. Chloramines thus formed are effective disinfectants and are
shown as combined available chlorine residual in figure (From A to B).
d. Further increase in the mole ratio of chlorine to ammonia result in formation of some
trichloramine and oxidation of part of ammonia to N2 and NO3-. These reactions are
essentially complete when 1.5 mole of chlorine has been added for each mole of ammonia
nitrogen originally present in water [i.e., C12:NH3-N = 1.5:1]. This is represented by the
portion of the curve from B to C.
e. Addition of chlorine beyond point C would produce free chlorine residuals and is referred
to as "breakpoint chlorination". In other words, chlorination of water to the extent that all
ammonia is converted to N2 or higher oxidation state is referred to as "breakpoint
chlorination'.

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f. Addition of chlorine beyond point C would produce free chlorine residuals and is referredto
as "breakpoint chlorination". In other words, chlorination of water to the extent that all
ammonia is converted to N2 or higher oxidation state is referred to as "breakpoint
chlorination".
APPARATUS:
1. Erlemeyer flask (250 mL)
2. Bottle
3. Beaker (250 mL)
4. Measuring cylinder
5. Dropper
6. Stirrer
REAGENTS:
1. Starch Indicator
2. Standard 0.025 N Sodium thiosulfate
3. Potassium Iodine crystal
4. Concentrated Acetic Acid
5. Chlorine water
PROCEDURE:
1. Take 100ml of sample in a conical flask and add a pinch of potassium iodide.
2. Add 5ml of acetic acid and allow the reaction to complete.
3. Titrate the sample against 0.0025N of sodium thiosulphate solution until the yellow colour
disappears.
4. Add 1ml of starch solution, blue colour appears then continue the titration until the blue
colour disappears (A-B).
RESULT:
Residual chorine in the given sample is………….mg/L
DISCUSSIONS:
Breakpoint chlorination is required to obtain a free chlorine residual for better disinfection if
ammonia is present in a water supply. While free chlorine residuals have good disinfecting
powers, they are usually dissipated quickly in the distribution system. For this reason, final
treatment with ammonia if often practiced to convert free chlorine residuals to longer-lasting
combined chlorine residuals. The difference between the amount of chlorine added to the
water and the amount of residual chlorine (i.e., free and combined available chlorine
remaining) at the end of a specified contact period is termed as "chlorine demand'.

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OBSERVATION TABLE:

CALCULATION:
Residual chlorine = ((A-B)×0.0025×35.45×1000)/ (volume of sample).

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DETERMINATION OF AVAILABLE CHLORINE IN BLEACHING

POWDER

Exp. No.: 11

AIM:
To determine the available chlorine in the given bleaching powder
THEORY
Bleaching powder is commonly used as a disinfectant. The chlorine present in the bleaching
powder gets reduced with time. So, to find the exact quantity of bleaching powder required,
the amount of available chlorine in the sample must be found out.

Chlorine will liberate free iodine from potassium iodide solution when its pH is 8 or less. The
iodine liberated, which is equivalent to the amount of active chlorine, is titrated with standard
sodium thiosulphate solution using starch as indicator.
APPARATUS:
Conical flask, Burette, Pipette
REAGENTS:

PROCEDURE:
1. Dissolve 1g bleaching powder in 1 litre of distilled water in a volumetric flask, and
stopperthe container.
(This can be done by first making a paste of the bleaching powder with mortar and
pestle.)
2. Place 5 mL acetic acid in an Erlenmeyer flask and add about 1g potassium iodide
crystals. Pour 25 mL of bleaching powder solution prepared above and mix with a
stirring rod.
3. Titrate with 0.025 N sodium thiosulphate solution until a pale yellow colour is
obtained. (Deep yellow changes to pale yellow.)
4. Add 1mL of starch solution and titrate until the blue colour disappears.
5. Note down the volume of sodium thiosulphate solution added (V1).
6. Take a volume of distilled water corresponding to the sample used.
7. Add 5 mL acetic acid, 1g potassium iodide and 1 mL starch solution.
8. If blue colour occurs, titrate with 0.025 N sodium thiosulphate solution until the blue
colour disappears.
9. Record the volume of sodium thiosulphate solution added (A1).

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10. If no blue colour occurs, titrate with 0.025 N iodine solution until a blue colour
appears. Note down the volume of iodine (A2).
11. Then, titrate with 0.025 N sodium thiosulphate solution till the blue colour disappears.
Record the volume of sodium thiosulphate solution added (A 3). Note down the
difference between the volume of iodine solution and sodium thiosulphate as
A4(A4=A2- A3).

RESULTS:
Available chlorine in the given bleaching powder is ______%

DISCUSSIONS:

The determination of available chlorine in a sample of bleaching powder is that a good

quality sample typically contains 35-38% of available chlorine. This percentage can decrease
over time due to auto-oxidation, making the determination crucial for assessing the
effectiveness of bleaching powder as a disinfectant or bleaching agent.

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OBSERVATIONS AND CALCULATIONS:

Bleaching powder solution x Standard sodium thiosulphate solution (0.025 N)

No. Volume of Bleaching Burette reading Burette reading Volume in mL

Powder in mL Initial Final

Distilled water × Standard sodium thiosulphate solution (0.025 N)

No. Volume of Bleaching Burette reading Burette reading Volume in mL

Powder in mL Initial Final

Distilled water x Standard iodine solution (0.025N)

No. Volume of Bleaching Burette reading Burette reading Volume in mL

Powder in mL Initial Final

mg of Cl2/mL (B) =

1000 mL of bleaching powder solution contains 1000 x B mg of Cl 2

i.e., 1000 mg bleaching powder contains 1000 B mg of Cl2

therefore, 100 mg of bleaching powder contains =

% of chlorine available = __________

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DETERMINATION OF SULPHATES

Exp. No.: 12

AIM:
To determine the available sulphates in the given sample
THEORY

APPARATUS:

REAGENTS:

PROCEDURE:

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OBSERVATIONS AND CALCULATIONS:

RESULTS:

The available sulphates in the given sample……………..

DISCUSSIONS:

The presence and concentration of sulfates are significant in wastewater as they can be
reduced to hydrogen sulfide (H2S) under anaerobic conditions, leading to odor problems and
sewer corrosion. Determining sulfate levels helps in managing and treating wastewater
effectively to prevent these issues.

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DETERMINATION OF FLUORIDE

Exp. No.: 13

AIM:
To determine the available Fluoride in the given sample
THEORY
Fluoride ions have dual significant in water supplies. High concentration of Fcauses dental
fluorosis (disfigurement of the teeth). At the same time, a concentration less than 0.8mg/L
results in ‘dental caries’. Hence, it is essential to maintain the F- concentration between 0.8 to
1.0mg/L in drinking water. In cases when fluoride concentration is less than 0.6 mg/l ,
supplementation (addition) of fluoride is necessary. Accurate determination of fluoride has
increased the importance in public water supply system as a public health measure.
Maintenance of an optimal fluoride concentration is essential is maintaining effectiveness and
safety of the fluoridation procedure.
DETERMINATION
A. Preliminary Distillation
A fluoride can be separated from other non-volatile constituents in water by conversion to
hydrofluoric or fluorosilicic acid and subsequent distillation. The conversion is obtained by
using a strong, high boiling acid distillation will separate most of the interfering ions of
fluoride and thus cause separation of fluoride ions.
B. SPANDS method for determination of fluoride
Principle: The spands colourimetric method is based on reaction between the fluoride and a
zirconium dye. Fluoride reacts with the zirconium dye dissociating a portion of it into a
colourless complex. As the amount of fluoride increases the colour produced becomes
progressively lighter. Increasing fluoride concentration decreases the intensity of colour
compelx due to bleaching action of fluoride and here the Beer’s law is obeyed inversely.
REAGENTS:

1. Standard Fluoride solution: Dissolve 221 gm of anhydrous sodium fluoride in distilled

water and dilute to 1 litre. 1 ml = 100 µg of fluoride.
2. Working standard fluoride solution Take 100 ml from standard fluoride solution and dilute
to 1 litre. 1 ml = 10 µg of fluoride.
3. SPANDS reagent solution:
Dissolve 958 mg SPANDS, sodium 2-(parasulfophenylazo)-1,8-dihydroxy-3,6- naphtalene
disulfonate, also called 4,5-dihydroxy-3-(parasulfophenlazo)-2,7- naphthalenedisulfonic acid
trisodium salt, in distilled water and dilute to 500 ml.
4. Zirconyle acid reagent Dissolve 133 mg zirconyl chloride octahydrate, (ZrOCl2.8H2O) in

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about 25 ml distilled water. Add 350 ml conc. HCl and dilute to 500 ml with distilled water.
5. Reference Solution Add 10 ml of SPANDS solution to 100 ml distilled water. Dilute 7 ml
conc HCl to 10 ml and add to the diluted SPANDS solution. The resulting solution, used for
setting the instrument reference point (zero).
6. Sodium arsenite solution: Dissolve 5.0 gms Na AsO2 and dilute to 1 L with distilled water.
PROCEDURE:
Take standards of 0,10,20,30 and 40 ml of standard fluoride solution (1ml = 10 µg). Add 10
ml zicronyle reagent. Take sample volume of 50 ml. add 10 ml SPAND’s reagent to all tubes
for colour development, mix well and read absorption at 575 nm. Set the 0 with reference
solution.
OBSERVATIONS AND CALCULATIONS:

Consentration (µg) Absorption at 575 nm

10
20
30
40
50

RESULTS:
The available Fluoride in the given sample………………

DISCUSSIONS:

Based on health considerations, the concentration of fluoride in drinking water should not
exceed 1.5 mg/L. Fluoride occurs naturally in seawater (1.4 mg/L), soil (up to 300 parts per
million) and air (from volcanic gases and industrial pollution).

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DETERMINATION OF DISSOLVED OXYGEN

Exp. No.: 14

AIM:
To determine dissolved oxygen of given sample.
THEORY
DO rapidly oxidizes the divalent manganous to its higher valency which forms a brown
hydrated oxide precipitate after addition of NaOH and KI. In the presence of iodide ions in an
acidic solution the oxidized manganese reverts to the divalent state and liberates Iodine from
KI equivalent to the original DO content. The liberated Iodine is then titrated against Sodium
thiosulphate solution with starch as an indicator. MnSO4 reacts with alkali to form white
precipitate Mn(OH)2 thus indicating absence of oxygen in the sample
APPARATUS:
BOD bottles (capacity 300 ml), burette, pipettes, conical flask, burette stands, tile, measuring
cylinder, weight balance, glass rod, beakers.
REAGENTS:
1. Winkler’s A solution:
Dissolve 48 gm tetra hydrate manganous sulphate in 100 ml distilled water. Filter if
necessary.
2. Winkler’s B solution:
Dissolve 50 gm of NaOH and 15 gm of KI in 100 ml distilled water.
3. Starch indicator:
Make a paste of 2gm L.R grade soluble starch powder in distilled water. Pour this
solution in 100 ml distilledwater. Boil for few minutes. Cool the solution and then
use.
4. Concentrated sulphuric acid
5. Standard sodium thiosulphate solution:
Dissolve 24.82 gm Na2S2O3 in distilled water and make up to 1 liter. It becomes 0.1N.
Take 250 ml of thissolution and make up to 1 liter with distilled water to prepare 0.025N.

PROCEDURE:
1. Collect the sample in 300ml BOD bottle.
2. Add 2ml of Winkler’s A solution and 2ml of Winkler’s B solution well below the surface
through the walls.
3. Stopper immediately to remove air bubbles and mix carefully by inverting bottle up and
down.

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4. Allow the brown precipitate to settle down leaving clear supernatant.

5. Add conc. Sulphuric acid drop by drop till precipitate digested.
6. Restopper the bottle and mix by inverting several times for complete dissolution.
7. A yellow colored solution appears.
8. Take 50 ml samples in conical flask.
9. Add few drops of starch indicator and titrate against 0.025N Na2S2O3 solution.
10. Note down the reading until the color changes from blue to colorless.
REACTION:

RESULT:
The amount of dissolved oxygen determined from the given sample is------- mg/l
DISCUSSIONS:
1. A minimum DO of 4 to 6 mg/l is desirable for the survival of aquatic life; higher values of
DO may cause corrosion of iron and steel.
2. High temperature, biological impurities, ammonia, nitrites, ferrous iron, chemicals such as
hydrogen sulphide and organic matter reduces DO values.
3. Drinking water should be rich in DO for good taste.

APPLICATION OF DATA:
1. It is necessary to know DO levels to assess the quality of raw water and to check on stream
pollution.
2. DO test is basis for BOD test which is an important parameter to evaluate organic pollution
potential of waste.
3. DO test is used to control oxygen in boiler feed water.
4. DO test is used to evaluate the pollution strength of domestic and industrial waste.
5. Determination of DO in waste water is useful to identify the nature of biochemical
reactions whether aerobic or anaerobic.

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OBSERVATIONS AND CALCULATIONS:

1. In burette: 0.025 N sodium thiosulphate solution.

2. In conical flask: 50 ml sample + indicator.
3. Indicator: starch.
4. End point: blue to colorless.

Sl No Burette reading Difference Average

DO (mg/l) = (V x 70) / u
Where,
V= ml of titrant used for DO determination (B.R.)
u = ml of water sample taken
70 = correction factor

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DETERMINATION OF NITRATE

Exp. No.: 15

AIM:
To determine the available Nitrate in the given sample
THEORY
Nitrate is determined by measuring the absorbance at 220nm in sample containing 1mL of
hydrochloric acid (1N) in 100mL sample. The concentration is calculated from graph from
standard nitrate solution in range 1-11mg/L as N
APPARATUS:
a. Spectrophotometer, for use at 220nm and 275nm with matched silica cells of 1cm or longer
light path.
b. Filter: One of the following is required.
i) Membrane filter: 0.45µm membrane filter and appropriate filter assemble
ii) Paper: Acid-washed, ashless hard-finish filter paper sufficiently retentive for fine
precipitates.
c. Nessler tubes, 50mL, short form
REAGENTS:
a. Redistilled water: use redistilled water for the preparation of all solutions and dilutions. b.
Stock nitrate solution: dissolve 721.8mg anhydrous potassium and dilute to 1000 ml with
distilled water. 1mL = 100 µg N = 443µg NO3-.
c. Standard nitrate solution: dilute 100mL stock nitrate solution to 1000mL with distilled
water. 1mL = 10µg NO3 N = 44.3µg NO3.
d. Hydrochloric acid solution: HCl, 1N.
e. Aluminium hydroxide suspension: dissolve 125g potash alum in 1000mL distilled water.
Warm to 60°C, add 55-60mL NH4OH and allow to stand for 1h. Decant the supernatant and
wash the precipitate a number of times till it is free from Cl, NO2 and NO3. Finally after
setting, decant off as much clean liquid as possible, leaving only the concentrated suspension.
CALIBRATION:
Prepare nitrate calibration standards in the range 0 to 350µg N by diluting 1, 2, 4, 7…..35mL
of the standard nitrate solution to 50mL. Treat the nitrate standards in the same manner as the
samples.
UV spectrophotometer method
The method is useful for the water free from organic contaminants and is most suitable for
drinking. Measurement of the ultraviolet absorption at 220nm enables rapid determination of

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nitrate. The nitrate calibration curve follows Beer’s law upto 11mg/L N. Acidification with
1N hydrochloric acid is designed to present interference from hydroxide or concentrations up
to 1,000mg/L as CaCO. Chloride has no effect on the determination. Minimum detectable
concentration is 40µg/L NO3- N.
PROCEDURE:
Read the absorbance or transmittance against redistilled water set at zero absorbance or 100%
transmittance. Use a wavelength of 220 nm to obtain the nitrate reading and, if necessary, a
wavelength of 275nm to obtain interference due to dissolved organic matter.

CALCULATION:
For correction for dissolved organic matter, subtract 2 times the reading at 275nm from the
reading at 220nm to obtain the absorbance due to nitrate. Convert this absorbance value into
equivalent nitrate by reading the nitrate value from a standard calibration curve.
Nitrate N, mg/L = mg nitrate-N /mL of sample NO3, mg/L = Nitrate N mg/L x 4.43

RESULT:
The amount of Nitrate determined from the given sample is------- mg/l
DISCUSSIONS:
The nitrate concentration in surface water is normally low (0–18 mg/L; up to ~4 mg/L as
nitrate-nitrogen), but can reach high levels as a result of agricultural runoff, refuse dump
runoff or contamination with human or animal wastes.

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DETERMINATION OF PHOSPHATE

Exp. No.: 16

AIM:
To determine the available Phosphate in the given sample
THEORY:
In a dilute orthophosphate solution, ammonium molybdate reacts under acid conditions to
form the heteropoly acid, molybdophosphoric acid. In the presence of vanadium, yellow
vanadomolybdo-phosphoric acid is formed. The intensity of the yellow color is proportional
to phosphate concentration. Absorbance may be measured at 400, 420 or 470 nm depending
on the sensitivity required, the absorbance at 400 nm being ten times that at 490 nm for a
given P concentration. However, due to potential interference from ferric ions at low
wavelengths, 470 nm is normally chosen. In this experiment, you will determine the
phosphorus content of various samples of waters. No sample pre-treatment is required, except
for dilution and addition of the colorimetric reagent. To avoid any possible phosphate
contamination from detergents, it is very important that you use only the acid-washed
glassware supplied in the cupboard for this experiment.
APPARATUS:
UV/visible spectrophotometer & glass cells
Volumetric flasks: 13 x 50 ml
Beakers: 6 x 100 ml
Pipettes: 10 & 25 ml bulb pipettes
REAGENTS:
Sample of water from the lungs of the victim.
Samples of water from the bath, swimming pool, tap, river and sea.
Vanadate-molybdate reagent: a mixture of ammonium molybdate [(NH4)6Mo7O24] and
ammonium metavanadate [NH4VO3] in HCl.
Standard phosphate solution: KH2PO4 containing 50.0 !g P per ml.
PREPARATION OF CALIBRATION STANDARDS:
Prepare a series of calibration standards by transferring 0, 5, 10, 15 and 20 ml aliquots of the
standard phosphate solution to 50 ml volumetric flasks, adding 10 ml of the vanadate-
molybdate reagent to each and diluting to the mark with deionized water. The standards thus
diluted contain 0, 5, 10, 15 and 20 !g P per ml (the one containing no phosphate acts as a
reagent blank).

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PROCEDURE:
1. Shake the ‘bath water sample’ to ensure thorough mixing, then transfer a 25-ml
aliquot to each of two 50-ml volumetric flasks.
2. Add 10 ml of the vanadate-molybdate reagent to one of these flasks - the second acts
as a sample blank - and dilute each to the mark with de-ionised water.
3. Shake to mix.
4. Repeat 1-3 for the other water samples.
5. Leave all prepared solutions (including calibration standards) to stand for at least 10
minutes for the color to develop fully before taking absorbance readings.
6. Measure the absorbance of each standard, sample and blank at 470 nm.
7. Plot a calibration graph of absorbance vs. phosphorus concentration (in !g/ml) for the
standard solutions.
8. Use this graph to determine the phosphate (as P) concentration in each water sample
(remember to allow for the dilution of the sample in your calculations) and subtract
the sample blank reading from the corresponding sample.

RESULT:
The amount of Phosphate determined from the given sample is------- mg/l
DISCUSSIONS:
Recommended phosphate levels in water vary depending on the water body and its intended
use, with lower concentrations generally considered better to prevent eutrophication and
maintain water quality. The U.S. Environmental Protection Agency (EPA) suggests a
maximum total phosphorus concentration of 100 µg/L (0.1 mg/L) in rivers and streams and
25 µg/L (0.025 mg/L) in lakes. Higher levels, even as low as 5 µg/L (0.005 mg/L) in some
estuaries, can cause eutrophication.

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DETERMINATION OF ARSENIC IN WATER

Exp. No.: 17

AIM:
To determine the ARSENIC content in the given sample
THEORY:
Arsenic is a major environmental pollutant and exposure occurs through environmental,
occupational and medicinal sources. Airborne exposure is small except in polluted locations.
Food exposure can be significant but, particularly in fish and shellfish, it is mostly in organic
forms that are relatively nontoxic. Drinking water remains the most significant source
worldwide, and large numbers of people are subject to serious exposure from this source.
Toxicity consists mostly of neuropathy, skin lesions, vascular damage, and carcinogenesis.
Vascular lesions are the result of endarteritis (blackfoot disease). This appears to be more
prevalent in developing rather than developed countries and may be related to nutritional
deficiencies. Skin cancer is the most clearly associated malignancy related to arsenic
exposure from drinking water; however, bladder, lung, liver, and kidney tumors also appear
to be related. There is no particular remedial action for chronic arsenic poisoning. Low
socioeconomic status and malnutrition may increase the risk of chronic toxicity.
ANALYTICAL METHODS FOR MEASURING ARSENIC:
The most commonly used method for detection of arsenic concentration water may be
categorized as follows:
1. Inductively coupled plasma (ICP) method
2. Hydride generation atomic absorption spectrophotometric method
3. Graphite furnace atomic absorption spectrophotometric method
4. Hydride generation-scraper-spectrophotometric (SDOC) method
5. Hydride generation-scraper-indicator paper-field kit
The first three methods involve high-cost equipment and provide more accuracy and lower
detection limit (minimum detection limit, MDL = 1 μg/L). The last two methods are
relatively low-cost methods but accuracy of determination is less.
Inductively coupled plasma (ICP) method
An ICP source consists of a flowing stream of argon gas ionized by applied radio frequency
field typically oscillating at 27.1 MHz. The water sample is atomized at temperature about
6000 to 8000.0 K. The light emitted from ICP Hydride generation atomic absorption
spectrophotometric method is focused on entrance slit and using radio frequency determines
absorbance of arsenic.
Hydride generation atomic absorption spectrophotometric method
In this method arsenic is reduced to gaseous arsine in a reaction vessel. The method is two

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types: i) manual hydride generation and ii) continuous hydride generation. In manual method
zinc is added to speed the reaction whereas continuous in continuous hydride generation no
zinc is needed. In continues measurement hydride generator a peristaltic pump is used to
meter and mix reagents and a gas-liquid separator unit uses flow of argon to strip out
hydrogen and arsine gas.
Hydride generation-sera per-spectrophotometric (SDDC) method
Minimum detectable quantity for this method is 1 micro gram As. This method essentially
involves: conversion (reduction) of all arsenic in water into As(III) and generation of arsine
gas in the form of arsenic hydride (AsH3). Absorbance of red-coloured complex produced by
passing of arsine gas through a solution of silver diethyl-dithiocarbamate (SODC) is
measured in a spectrophotometer at 535 nm wavelength.
Hydride generation-scraper-indicator paper-field kit
A simple and reasonably accurate method for arsenic measurement Similar to SOOC method
all arsenic in water is converted to As(III) and generates arsine gas which is then passes
through a filter paper soaked in mercuric bromide.
Laboratory Measurement of Arsenic by Arsenic Tool kit Method:
In the class arsenic measurement by arsenic tool kits will be conducted. The kit involves the
generation of arsine (AsH3) from inorganic arsenic species by reduction with Zn and HCl.
The arsine then reacts with a test strip containing HgBr2 to produce a color that is compared
with a color scale for quantitation.
APPARATUS:
1. Arsenic toolkits
PROCEDURE:
1. Add reagent to Reaction Bottle and shake vigorously.
2. Insert the strip into the turret and close it. Let it sit 10 minutes.
3. Select the As concentration on the chart that matched the color of the test strip most
closely. The reference chart provided with the kit displays the yellow to brown range of
colors expected for As concentrations of 0, 10, 25, 50, 100, 200, 300, 500, and 1000 μg/L.

OBSERVATIONS:

Sample No Source of Sample Temperature of Arsenic

Sample (°C) concentration (μg/L)

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RESULTS:
Arsenic concentration in the given sample ………………………(μg/L)
DISCUSSIONS:
According to BECR 2023, drinking water standard for arsenic in Bangladesh is 50 μg/L (or
0.05 mg/L). The WHO guideline value for arsenic in drinking water is 10μg/Land the
USEPA is also planning to revise its standard from 50 μg/L to 10μg/L.

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DETERMINATION OF TOTAL AND FECAL COLIFORM IN WATER

Exp. No.: 18

AIM:
To determine the total and fecal coliform content in the given sample
THEORY:
Total coliforms are grouped into two categories (1) Fecal coliform (thermo-tolerant
coliform-) and (2) non-fecal coliform. Total coliforms are defined as gram negative bacteria
which ferment lactose at 35° or 37° C with the production of acid, gas and aldehyde within
24 or 48 hours. Fecal coliform is a subgroup of total coliforms, which live in the warm-
blooded animals and have the same properties as the total coliform but tolerate and grow at
higher selective temperature range of 44° to 44. 5°C.In addition, they form indole from
tryptophan. And these combined properties, when positive, are regarded as presumptive
Escherichia coli (presumptive E. coli). Some coliform species are frequently associated with
plant debris or may be common inhabitants in soil or surface waters which arc called non-
fecal coliforms.
Total coliform (TC) = Fecal coliform (FC) + Non-fecal coliform
APPARATUS:
1. Petri Dish
2. Incubator
3. Measuring Cylinder, beaker, dropper etc.
REAGENTS:
1. Appropriate culture medium (broth)
2. Distilled water
PROCEDURE (FOR MEMBRANE FILTER (MF) METHOD):
Determination of Total Coliforms (TC):
1. Connect the Erlenmeyer (side-arm) flask to the vacuum source (turned off) and place the
porous support in position. if an electric pump is used, it is advisable to put a second flask
between the Erlenmeyer and the vacuum source; this second flask acts as a water trap and
thus protects the electric pump.
2. Open a Petri-dish and place a pad in it.
3. 'With a sterile pipette add 2 mL of selective broth (culture) medium to saturate the pad.
4. Assemble the filtration unit by placing sterile membrane filter on the porous support, using
forceps sterilized earlier by flaming.
5. Place the upper container in position and secure it with the special clamps. The type of
clamping to be used will depend on the type of equipment.

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6. Pour tide volume of sample chosen as optimal, in accordance with the type of water, into
the upper container. If the test sample is less than 10 mL, at least 20 ml of sterile dilution
water should be added to the top container before filtration applying the vacuum.
7. After the sample has passed through the filter, disconnect the vacuum and rinse the
container with 20-30 mL of sterile dilution water. Repeat the rinsing after all the water from
the first rinse has passed through the filter.
8. Take the filtration unit apart and using the forceps, place the membrane filter in the Petri-
dish on the pad with the grid side up. Make sure that no air bubbles are trapped between the
pad and the filter.
9. Invert the Petri-dish for incubation.
10. Incubate at 35°C or 37°C for 18-24 hours with 100% humidity (to ensure this, place a
piece of wet cotton wool in the incubator). If ointment containers or plastic dishes with tight-
fitting lids are used, humidification is not necessary.
Bacterial Colony observation:
Colonies of coliform bacteria are a medium red or dark red color, with a greenish gold or
metallic surface sheen. This sheen may cover the entire colony or appear only in the centre of
the colony. Colonies of other types should not be counted. The colonies can be counted with
the aid of a lens. The number of total coliforms per 100 mL is then given by:
Determination of Fecal Coliforms (FC):
The procedure for fecal coliforms is similar to that used for determining total coliforms. Filter
the sample as described, and place the membrane filler on the pad saturated with appropriate
culture medium.
1. Place the dishes in an incubator at 44±0.5 °C for 24 hours at 100% humidity. Alternatively,
tight-fitting or sealed Petri-dishes may be placed in water-proof plastic bags for incubation.
2. Submerge the bags in a water-bath maintained at 44±0.5°C for 24 hours. The plastic bags
must be below the surface of the water throughout the incubation period. They can be held
down by means of a suitable weight, e.g., a metal rack.
Bacterial Colony observation:
Colonies of fecal coliform bacteria are blue in color. This color may cover the entire colony,
or appear only in the center of the colony. Colonies of other types should not be counted. The
colonies can be counted with the aid of a lens. The number of fecal coliforms per 100 ml is
then given by:
OBSERVATIONS AND CALCULATIONS:

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Observation Total Coliform Unit per 100 ml Fecal Coliform Unit per
No. 100 ml

RESULTS:
Total Coliform in the given sample ………………………
Fecal Coliform in the given sample ………………………
DISCUSSIONS:
The measurement of total coliforms is of particular relevance for treated and / or chlorinated
water supplies; in this case the absence of total coliforms would normally indicate that the
water has been sufficiently treated / disinfected to destroy various pathogens. Measurement
of focal coliforms is a better indicator of general contamination by material of fecal origin.
The predominant species of fecal coliform group is Escherichia coil (E. coil), which is
exclusively of fecal origin, but strains of Klebsella pneumonia and Enterobacter species may
also be present in contaminated water.

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REFERENCES

1. Standard Methods for the Examination of Water and Wastewater, 23rd edition,
American Public Health Association, American Water Works Association,
Water Environment Federation, 2017.
2. Water Supply Engineering, 33rd edition, Santhosh Kumar Garg, Khanna
publishers.
3. Sewage Disposal and Air Pollution Engineering, 39th edition, Santhosh Kumar
Garg, Khanna publishers.
4. IS: 10500:2012 Drinking Water - Specification, Second revision, Bureau of
Indian Standards, 2012.

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