PREFACE

This report is prepared by the student of Environmental Engineering, UET Taxila and is
submitted to Engr. Arfa Iqbal
This is a detailed on report on different experiments done in the the lab. It includes
experiments based on spectrophotometer to check maximum wavelength and to find
unknown concentrations of samples. It also includes methods to check parameters of water
like metal concentrations and oil and grease content.
Above all I would like to thank Almighty Allah for enabling us to accomplish our work and
would like to thank Ma’am Arfa Iqbal for her cooperation and assistance.
.

P a g e 1 | 34
Contents
LAYOUT OF ENVIRONMENTAL ANALYTICAL TECHNIQUES LAB............................5
DESCRIPTION OF APPARATUS:.......................................................................................6
EXPERIMENT NO. 1..............................................................................................................10
TO DETERMINE λmax OF GIVEN SAMPLE BY SPECTROPHOTOMETER.....................10
1.1 THEORY:.......................................................................................................................10
1.1 1 Wavelength:.............................................................................................................10
1.1.2 Spectroscopy:..........................................................................................................10
1.1.3 Spectrophotometer:..................................................................................................10
1.1.4 Parts of spectrophotometer:.....................................................................................11
1.1.5 Working of spectrophotometer:...............................................................................11
1.2 APPARARTUS:.............................................................................................................11
1.3 PROCEDURE:...............................................................................................................11
1.4 OBSERVATIONS AND CALCULATIONS:...............................................................11
1.4.1 For K2CrO4...............................................................................................................11
1.4.2 For K2Cr2O7..............................................................................................................12
1.4.3 For KMnO4..............................................................................................................13
1.5 COMMENTS.................................................................................................................14
EXPERIMENT NO. 2..............................................................................................................15
ESTIMATION OF COPPER ION CONCENTRATION IN GIVEN SOLUTION BY UV-
SPECTROPHOTOMETER.....................................................................................................15
2.1 THEORY:.......................................................................................................................15
2.1.1 Copper:.....................................................................................................................15
2.1.2 Environmental Significance of Copper:..................................................................15
2.1.3 Guidelines for Copper Concentration:.....................................................................16
2.2 APPARATUS:................................................................................................................16
2.3 PROCEDURE:...............................................................................................................16
2.4 OBSERVATIONS AND CALCULATIONS:...............................................................17
EXPERIMENT NO. 3..............................................................................................................19
ESTIMATION OF IRON BY UV SPECTROPHOTOMETER..............................................19
3.1 THEORY:.......................................................................................................................19
3.1.1 Iron:..........................................................................................................................19
3.1.2 Occurrence:..............................................................................................................19

P a g e 2 | 34
 3.1.3 Major Uses:..............................................................................................................19
3.1.4 Guidelines and Standards:.......................................................................................19
3.1.5 Excess of iron...........................................................................................................20
3.1.6 Iron Removal Techniques:.......................................................................................20
3.2 APPARATUS:................................................................................................................20
3.3 PROCEDURE:...............................................................................................................20
3.4 OBSERVATIONS AND CALCULATIONS:...............................................................21
3.5 COMMENTS:................................................................................................................21
EXPERIMENT NO. 4..............................................................................................................22
ESTIMATION OF Ni IN WATER SAMPLE.........................................................................22
4.1 THEORY:.......................................................................................................................22
4.1.1 Nickel:......................................................................................................................22
4.1.2 Occurrence of nickel:...............................................................................................22
4.1.3 Environmental Significance of Nickel:....................................................................22
4.1.4 Guidelines for Nickel Concentration:......................................................................22
4.2 APPARATUS:................................................................................................................22
4.3 PROCEDURE:...............................................................................................................23
4.4 OBSERVATION AND CALCULATIONS:.................................................................23
4.5 COMMENTS:................................................................................................................24
EXPERIMENT NO. 5..............................................................................................................25
ESTIMATION OF OIL AND GREASE CONTENT..............................................................25
5.1 THEORY:.......................................................................................................................25
5.1.1 Introduction:.............................................................................................................25
5.1.2 Oil and grease in waste water:.................................................................................25
5.1.3 Forms of Existence:.................................................................................................25
5.1.4 Environmental Significance:....................................................................................25
5.1.5 Removal Techniques:..............................................................................................26
5.2 APPARATUS:................................................................................................................26
5.3 PROCEDURE:...............................................................................................................26
5.4 OBSERVATIONS:.........................................................................................................26
5.5 COMMENTS:................................................................................................................26
EXPERIMENT NO. 6..............................................................................................................27
ESTIMATION OF NITROGEN CONTENT IN WATER......................................................27

P a g e 3 | 34
 6.1 THEORY:.......................................................................................................................27
6.1.1 Introduction:.............................................................................................................27
6.1.2Measuring Nitrogen:.................................................................................................27
6.1.3Why Nitrogen measurement is necessary?...............................................................27
6.1.4 Environmental Significance:....................................................................................28
6.2 APPARATUS:................................................................................................................28
6.3 PROCEDURE:...............................................................................................................28
6.3 OBSERVATION AND CALCULATION.....................................................................28
6.4 COMMENTS:................................................................................................................29
STUDY OF INSTRUMENTS.................................................................................................29
8.1 ATOMIC ABSORBANCE SPECTROPHOTOMETER (AAS)...................................29
8.1.1Atomic Absorption Spectroscopy (AAS).................................................................30
8.2 TOTAL ORGANIC CARBON ANALYZER................................................................33

P a g e 4 | 34
LAYOUT OF ENVIRONMENTAL ANALYTICAL TECHNIQUES LAB

P a g e 5 | 34
DESCRIPTION OF APPARATUS:
1- Titration Apparatus:

Figure 1 Titration Apparatus

The technique of titration is used to find out accurately how much of a chemical
substance is dissolved in a given volume of a solution, that is, the concentration of the
solution.
It consists of a burette to measure volume of solution added, pipette which is used to
make an accurate volume of solution and volumetric flask to make up standard
solution.
2- Imhoff Cone:

Figure 2 Imhoff Cone

A clear, cone-shaped container marked with graduations. The cone is used to measure
the volume of settleable solids in a specific volume (usually one liter) of water or
wastewater.
3- Column Test Apparatus:

Figure 3 Column Test Apparatus

It is used to measure different charcteristics of water e.g the settling characteristics.

P a g e 6 | 34
4- TOC Analyzer:

Figure 4 TOC Analyzer

It is used to measure total organic carbon content in water and waste water.

5- Weighing Balance:

Figure 5 Weighing Balance

It is used to measure mass of samples.

6- Desiccator:

Figure 6 Desiccator

Desiccators are sealable enclosures containing desiccants used for preserving

moisture-sensitive items such as cobalt chloride paper for another use. They are also
used to remove moisture from any sample.

7- Nanofiltration Apparatus:
P a g e 7 | 34
 Figure 7 Nanofiltration Apparatus

It is used for water purification.  Nanofiltration (NF) employs a pressure gradient to

selectively transport solvent and certain solutes across a membrane

8- Oil and Greese Analyzer:

Figure 8 Oil and Greese Analyzer

It is used to measure oil and grease content in water and waste water sample.

9- Water Sampler:

Figure 9 Water Sampler

It is used to collect water samples at different depths.

10- Fridge:

P a g e 8 | 34
 Figure 10 Fridge

This is used to cool samples or specimens for preservation.

11- Spectrophotometer:

Figure 11 Spectrophotometer

 Spectrophotometer is the quantitative measurement of the absorption or transmission

properties of a material as a function of wavelength.

12- Jar Test Apparatus:

Figure 12 Jar Test Apparatus

The purpose of this test is to select types of coagulant (alum) and also to estimate the
optimal dose needed in removing the suspended particles that occurred in raw water.
Jar test is an experiment to understand the processes of coagulation, flocculation and
sedimentation.

P a g e 9 | 34
EXPERIMENT NO. 1

TO DETERMINE λmax OF GIVEN SAMPLE BY SPECTROPHOTOMETER

1.1 THEORY:
1.1 1 Wavelength:
Wavelength is defined as the distance between two
successive crests or troughs of wave.
This "visible light" corresponds to a wavelength range of 400 -
700 nanometres (nm) and a colour range of violet through red.
The human eye is not capable of seeing radiation
with wavelengths outside the visible spectrum. The visible
colours from shortest to longest wavelength are: violet, blue,
green, yellow, orange, and red.
1.1.2 Spectroscopy: Figure 1.1 Electromagnetic Spectrum

The study of molecular structure of any compound through absorbance, transmitting and
scattering of light is called spectroscopy.
 Absorbance:
Excitation of electrons i.e. transition of electrons from lower energy level to higher energy
level.
 Transmission:
De-excitation of electrons i.e. transition of electrons from higher energy level to lower energy
level.
1.1.3 Spectrophotometer:
A device which is used to measure absorbance, transmittance in terms of wavelength of light.
The most common spectrophotometers are used in the UV and visible regions of spectrum,
and some of these instruments also operate into the near infrared region as well.
Table 1.1 Wavelengths of different electromagnetic waves

Type of electromagnetic waves Wavelength (nm)

UV 200-400
Visible 400-700
Infrared Above 700
1.1.4 Parts of spectrophotometer:
 Spectrometer:

P a g e 10 | 34
It allows light of specific wavelength to pass.
 Photometer:
It detects the light that is absorbed or transmitted by sample.
1.1.5 Working of spectrophotometer:
 The light source shines through the sample.
 The sample absorbs the light.
 The detector detects how much light the sample has absorbed
 The detector then converts how much light the sample absorbed in number.
 The numbers are either plotted straight away or transmitted to a computer to be
further manipulated.
1.2 APPARARTUS:
Spectrophotometer
1.3 PROCEDURE:
i. Take three 0.01% solutions of K2Cr2O7, KMnO4, K2Cr2O7
ii. Place them one by one in cell and run UV spectrophotometer.
iii. Select different wavelength and note the absorbance.
iv. Draw calibration curve for all the three samples.
v. Determine λmax for each sample at maximum absorption from the graphs.
1.4 OBSERVATIONS AND CALCULATIONS:

1.4.1 For K2CrO4

Table 1.2 Absorption at different wavelngths for K2CrO4

Wavelength (nm) Absorption (Abs)

450 0.127
475 0.057
500 0.021
525 0.005
550 0.003
575 0.003
600 0.003

P a g e 11 | 34
 Graph 1.1 Wavelength vs Absorption for K2CrO4

Maximum absorption is at 450nm

1.4.2 For K2Cr2O7

Table 1.3 Absorption at different wavelengths for K2Cr2O7

Wavelength (nm) Absorption (Abs)

450 0.186
475 0.107
500 0.040
525 0.010
550 0.003

Graph 1.2 Wavelength vs Absorption for K2Cr2O7

P a g e 12 | 34
 Maximum absorption is at 450nm

1.4.3 For KMnO4

Table 1.3 Absorption at different wavelengths for KMnO4

Wavelength (nm) Absorption (Abs)

450 0.144
475 0.1480
500 1.195
525 1.913
550 1.680
575 0.772
600 0.216

Graph 1.3 Wavelength vs Absorption for KMnO4

Maximum absorption is at 525nm

1.5 COMMENTS
For K2CrO4 and K2Cr2O7 maximum absorption is at same wavelength which is 450nm. This
can be due to similarities in the molecular structure of both chemicals. For KMnO 4 it is
525nm due to its different characteristics e.g. molecular structure and frequency of movement
of molecules.

P a g e 13 | 34
Spectrophotometer works on the principle of Beer-Lambert law according to which
absorption is directly proportional to path length along which light moves and concentration
of sample. It is usually written as:
A=εlc

Path length for all the samples is same so the difference in absorption is due to the different
concentration of K2Cr2O7 and K2Cr2O7 in comparison to KMnO4.

P a g e 14 | 34
EXPERIMENT NO. 2

ESTIMATION OF COPPER ION CONCENTRATION IN GIVEN

SOLUTION BY UV-SPECTROPHOTOMETER

2.1 THEORY:
2.1.1 Copper:
Copper is a chemical element with
symbol Cu (from Latin: cuprum) and atomic
number 29. It is a soft, malleable and ductile metal
with very high thermal and electrical conductivity. A
freshly exposed surface of pure copper has a reddish-
orange colour. It is used as a conductor of heat and
electricity, as a building material, and as a constituent
of various metal alloys
It is a transition element and form cuprous (Cu +1) and Figure 2.1 Copper atom

cupric ions (Cu+2).

It is a chromophore (Those compounds having specific colour and visible in UV spectrometer
are called chromophores.)
It is found everywhere in water, soils, rocks, atmosphere etc.
2.1.2 Environmental Significance of Copper:
 Used in manufacturing of electric wires, pipes, coins etc.
 Used in food addictives
 Used in insecticides and fungicides
 Copper aides in formation of haemoglobin
 Copper helps iron in blood absorption
 Traditionally it has been one of the metals used to make coins, along with silver and
gold.
 Copper sulphate is used widely as an agricultural poison and as an algaecide in water
purification.
 Copper compounds, such as Fehling’s solution, are used in chemical tests for sugar
detection

P a g e 15 | 34
Excess of copper concentration in drinking water can cause vomiting, nausea, diarrhoea and
abdominal pain.
In drinking water colour of dissolved copper is pale blue. On porcelam (Koalin) sink, blue
green colour is due to copper.
Colour of anhydrous CuSO4 is pale blue green. It is also called blue stone or blue vitriol and
hydrated CuSO4.5H2O is dark blue. To measure copper concentration in sample anhydrous
copper is used this is because it gives colour.
2.1.3 Guidelines for Copper Concentration:
 NEQs/WHO guidelines for drinking water is 2mg/l
 NEQs for waste water is 1mg/l
2.2 APPARATUS:
i. Volumetric flask
ii. Beaker
iii. Spectrophotometer
2.3 PROCEDURE:
i. Prepare stock solution of copper (1000ppm) by adding 0.392g of CuSO 4.5H2O in
100ml of water.
ii. Prepare intermediate solution of copper (100ppm) by adding 10ml of stock solution
into a volumetric flask and diluting upto 100ml
iii. Prepare calibrated solutions of copper of the following concentrations:
(Add 10ml of NH4OH in each sample before adding water to enhance colour)
 20ppm: By adding 20ml of intermediate solution and adding upto 100ml of
water.
 40ppm: By adding 40ml of intermediate solution and adding upto 100ml of
water.
 60ppm: By adding 60ml of intermediate solution and adding upto 100ml of
water.
 80ppm: By adding 80ml of intermediate solution and adding upto 100ml of
water.
iv. Find absorbance of each calibrated sample at 620nm (wavelength at which maximum
absorbance of copper takes place).
v. Plot calibration curve between absorbance and concentration.
vi. Find absorbance for copper sample of unknown concentration.

P a g e 16 | 34
2.4 OBSERVATIONS AND CALCULATIONS:

Table 2.1 Absorbance at different concentration of copper

Concentration (ppm) Absorbance (Abs)

20 0.027
40 0.051
60 0.065
80 0.076
Unknown 0.039

Graph 2.1 Concentration vs Absorbance for copper

From the graph it can be seen that concentration at absorbance 0.039 is approximately 30 so,
Concentration of unknown sample is 30ppm

2.5 COMMENTS:
Using spectrophotometer is most convenient method to measure the unknown concentration
of copper but for this it is first necessary to establish relationship between concentration and
absorbance for copper solution. Then we can find unknown concentration from the
calibration curve.

P a g e 17 | 34
In this experiment, absorbance for sample of unknown concentration is 0.039 and from the
graph it is deduces that respective concentration is 30ppm or 30mg/L.

P a g e 18 | 34
EXPERIMENT NO. 3

ESTIMATION OF IRON BY UV SPECTROPHOTOMETER

3.1 THEORY:
3.1.1 Iron:
Iron is a chemical element with symbol Fe and atomic number 26. It is a metal in
the first transition series. Like other group 8 elements, iron exists in a wide range
of oxidation states, −2 to +6, although +2 and +3 are the most common. Elemental iron
occurs in meteoroids and other low oxygen environments, but is reactive to oxygen
and water. Fresh iron surfaces appear lustrous silvery-gray, but oxidize in normal air to
give hydrated iron oxides, commonly known as rust. Unlike many other metals which
form passivating oxide layers, iron oxides occupy more volume than the metal and thus
flake off, exposing fresh surfaces for corrosion.
Iron is the second most abundant metal on earth crust. It lies in the form Fe +2 (ferrous) and
Fe+3 (ferric). Ferric and ferrous combine with oxides, hydroxides and sulphates.
3.1.2 Occurrence:
Atmosphere, surface water, ground water, food e.tc
3.1.3 Major Uses:
 It is used in construction purposes
 It is used in water supply pipes
 Its compounds are used in pigments and paints
 Compounds of iron are used as coagulants for water and waste water treatment
 2/3rd of haemoglobin is made of iron compounds
3.1.4 Guidelines and Standards:
For drinking water
 WHO guideline is 0.2mg/l
 EPA guideline is 0.3mg/l
For waste water
 NEQs for iron is 2mg/l

P a g e 19 | 34
3.1.5 Excess of iron
 In drinking water it gives reddish brown colour, metallic and bitter taste and
odour problems.
 In water supply scheme excess of iron will cause sediment deposition, pressure
will be reduced so pump will require more energy to supply water.
 In laundry it will cause reddish brown colour on utensils and plumbing fixtures.
 Iron bacteria production will be caused. Bacteria will feed on iron and import
reddish brown colour.
3.1.6 Iron Removal Techniques:
Reverse Osmosis, Ultrafiltration, Nano filtration, Ion Exchange Method and Aeration.

3.2 APPARATUS:
i. Spectrophotometer
ii. Pipette
iii. Volumetric flask
3.3 PROCEDURE:
i. Prepare stock solution of ferrous ammonium sulphate hexahydrate of 100ppm by
adding 0.7g of Fe(NH4)2(SO4)2 in 10 to 15ml of distilled water in volumetric flask and
dissolve.
ii. Add concentrated H2SO4 drop wise till clear solution is obtained.
iii. Add 0.1N KMnO4 till you get faint pink colour.
iv. Half fill volumetric flask with water.
v. Dilute upto 100ml.
vi. Prepare calibrated solution of 10ppm, 20ppm, 30ppm and 40ppm
 For 10ppm add 1 ml of stock solution in volumetric flask
 For 20ppm add 2 ml of stock solution in volumetric flask
 For 30ppm add 3 ml of stock solution in volumetric flask
 For 40ppm add 4 ml of stock solution in volumetric flask
vii. Now add 0.5ml of concentrated HCl
viii. Add5-10ml hydroxylamine in flask
ix. Add 5ml ammonium acetate buffer
x. Add 10ml 1,10 phenonthrolene indicator
xi. Dilute upto 100ml

P a g e 20 | 34
xii. Place each sample in UV spectrophotometer and run at 510nm ( maximum
wavelength)
xiii. Plot graph between absorbance and concentration
xiv. Find absorbance of sample of unknown concentration and using calibration curve find
its concentration.
3.4 OBSERVATIONS AND CALCULATIONS:

Table 3.1 Absorbance at different concentration of Iron

Concentration (ppm) Absorbance (Abs)

10 1.411
20 1.612
30 1.865
40 2.606
Unknown 2.349

Table 3.1 Concentration vs Absorbance for Iron

3.5 COMMENTS:
Iron can form free radicals, its concentration in body tissues must be regulated
because in excessive amounts, it can lead to tissue damage. Disorders of iron metabolism are
among the most common diseases of humans and ranging from anaemia to iron overload, and
possibly to neurodegenerative diseases.
So this is a very convenient and simple method to find the concentration of copper.

P a g e 21 | 34
EXPERIMENT NO. 4

ESTIMATION OF Ni IN WATER SAMPLE

4.1 THEORY:
4.1.1 Nickel:
Nickel is a chemical element with symbol Ni and atomic number 28. It is a silvery-
white lustrous metal with a slight golden tinge. Nickel belongs to the transition metals and is
hard and ductile. Pure nickel shows a significant chemical activity that can be observed when
nickel is powdered to maximize the exposed surface area on which reactions can occur, but
larger pieces of the metal are slow to react with air at ambient conditions due to the formation
of a protective oxide surface. An iron–nickel mixture is thought to compose Earth's inner
core. Pure native nickel is found in tiny amounts, usually in rocks.
It is 24rth most abundant element in earth.
4.1.2 Occurrence of nickel:
 It is present in food (peas, ground nuts), water, air and soil.
 It is added in environment through incinerators and power plants
 It is added in surface water when in passes through rocks whereas in ground water it
is added through pipes and stainless steel.
4.1.3 Environmental Significance of Nickel:
The major uses of nickel are:
 It is used in manufacture of stainless steel and alloys.
 It is used in coin making.
 It is used as catalyst in nickel cadmium batteries.
 It is used in food supplements.
 It resists corrosion and is used to plate other metals to protect them.
 Finely divided nickel is used as a catalyst for hydrogenating vegetable oils.
 Nickel steel is used for armour plating. Other alloys of nickel are used in boat
propeller shafts and turbine blades.
4.1.4 Guidelines for Nickel Concentration:
 According to NEQs nickel concentration in drinking water is 0.02mg/L
 According to NEQs nickel concentration in waste water is 1mg/L
4.2 APPARATUS:
i. Volumetric flask

P a g e 22 | 34
 ii. Measuring flask
iii. Pipette
iv. Spectrophotometer
4.3 PROCEDURE:
i. Prepare stock solution by adding 2g of NiSO4.6H2O in 100ml of water. This will yield
6100ppm solution of Nickel.
ii. Prepare calibrated solution in 5ml, 10ml, 15ml and 20ml.
 In 10ml solution, concentration of nickel is 305ppm
 In 10ml solution, concentration of nickel is 610ppm
 In 10ml solution, concentration of nickel is 915ppm
 In 10ml solution, concentration of nickel is 1220ppm
iii. Add 10ml of NH4OH in flask.
iv. Place each sample in UV spectrophotometer and check absorbance at 585nm.
v. Draw calibration curve between concentration and absorbance.
vi. Take unknown sample and add 10ml of NH4OH in it.
vii. Check absorbance of unknown sample and find concentration from the calibrated
sample.
4.4 OBSERVATION AND CALCULATIONS:

Figure 4.1 Absorbance at different concentration of Nickel

Concentration (ppm) Absorbance (Abs)

305 0.044
610 0.099
915 0.307
1220 0.430
Unknown 0.074

P a g e 23 | 34
 Graph 4.1 Concentration vs Absorbance for Nickel

The concentration of unknown sample is 500ppm

4.5 COMMENTS:
Major effects of excess of nickel ion concentration are that is one of many
carcinogenic metals known to be an environmental and occupational pollutant. The larger
part of all nickel compounds that are released to the environment will adsorb to sediment or
soil particles and become immobile as a result. So it is very important to have information
about iron concentration and this is the most reliable method.

P a g e 24 | 34
EXPERIMENT NO. 5

ESTIMATION OF OIL AND GREASE CONTENT

5.1 THEORY:

5.1.1 Introduction:
Grease and is a semisolid lubricant. Grease generally consists of a soap emulsified
with mineral or vegetable oil. The characteristic feature of greases is that they possess a high
initial viscosity, which upon the application of shear, drops to give the effect of an oil-
lubricated bearing of approximately the same viscosity as the base oil used in the grease. This
change in viscosity is called shear thinning. Grease is sometimes used to describe lubricating
materials that are simply soft solids or high viscosity liquids, but these materials do not
exhibit the shear-thinning properties characteristic of the classical grease
5.1.2 Oil and grease in waste water:
It is found in domestic wastewaters & certain industrial wastes and it cause problems
during treatment and for ultimate disposal.
Oil and grease are poorly soluble in water and separate from aqueous phase.
Wastewaters from meat packing industry and slaughter houses are high in oil and grease
content. Such industries should apply preliminary treatment for the recovery of oil / grease
before discharge.
Grease accumulates into ‘grease balls’ unsightly appearance to the surface of final settling
tank.
5.1.3 Forms of Existence:
Oil and grease occurs in two forms:
 Emulsified
 Free
5.1.4 Environmental Significance:
 It is carcinogenic
 In treatment plants it can cause clogging
 It can cause film formation which can disturb secondary treatment. Film is formed
around colloidal or suspended particles which resist formation.

P a g e 25 | 34
5.1.5 Removal Techniques:
 Oil and grease traps
 Oil and grease skimming devices
5.2 APPARATUS:
Oil and grease content analyser
5.3 PROCEDURE:
i. Take sample of waste water in a beaker
ii. Dip the probe of oil and content analyzer in sample
iii. Make the reading zero of the analyzer
iv. Note the reading
5.4 OBSERVATIONS:
3.1ppm
The oil and grease content of this waste water sample is

5.5 COMMENTS:
The oil and grease content of this waste water sample is within permissible limit
which is 10ppm as per NEQs. It may interfere in the secondary treatment of water and its
quantity can be reduced by oil and grease skims or traps.

P a g e 26 | 34
EXPERIMENT NO. 6

ESTIMATION OF NITROGEN CONTENT IN WATER

6.1 THEORY:
6.1.1 Introduction:
A colourless tasteless odourless
element that as a diatomic gas is
relatively inert and constitutes 78
percent of the atmosphere and that is
a constituent of organic compounds
found in all living tissues.
6.1.2Measuring Nitrogen:
Figure 6.1 Apparatus to measure nitrogen content
The Kjeldahl method or
Kjeldahl digestion in analytical chemistry is a method for the quantitative determination of
organic nitrogen in chemical substances like ammonia developed by Johan Kjeldahl in 1883.
The method consists of heating a substance with sulphuric acid, which decomposes the
organic substance by oxidation to liberate the reduced nitrogen as ammonium sulphate. In
this step potassium sulphate is added to increase the boiling point of the medium (from 337
°C to 373 °C). Chemical decomposition of the sample is complete when the initially very
dark-coloured medium has become clear and colourless.
The solution is then distilled with a small quantity of sodium hydroxide, which converts the
ammonium salt to ammonia. The amount of ammonia present, and thus the amount of
nitrogen present in the sample, is determined by back titration. The end of the condenser is
dipped into a solution of boric acid. The ammonia reacts with the acid and the remainder of
the acid is then titrated with a sodium carbonate solution by way of a methyl orange pH
indicator.
6.1.3Why Nitrogen measurement is necessary?
Nitrogen (N) is one of the building blocks of life: it is essential for all plants and animals to
survive. Nitrogen (N2) makes up almost 80% of our atmosphere, but it is an unreactive form
that is not accessible to us.
Humans and most other species on earth require nitrogen in a “fixed,” reactive form. Reactive
nitrogen is necessary for the food production process. Until the discovery of the Haber-Bosch
process in the early 1900s, we only had access to naturally occurring sources of reactive
P a g e 27 | 34
nitrogen (such as manure and guano) for food production. The Haber-Bosch process--an
industrial process through which we can fix reactive nitrogen--has allowed food production to
keep up with the growing human population, but at a cost to the environment.
6.1.4 Environmental Significance:
1. Nitrogen is an important building block of proteins, nucleic acids and other cellular
constituents which are essential for all forms of life. Nitrogen is such an important key
nutrient element for plants that it warrants careful management, and – if mismanaged –
can lead to severe environmental problems.
2. Excess nitrogen can cause Algae and aquatic plants to grow too rapidly. This growth may
be good for those life forms, but it may cause problems for others

6.2 APPARATUS:
i. Volumetric flask
ii. Measuring flask
iii. Pipette
iv. Distillation assembly

6.3 PROCEDURE:
1- Take 140 ml of waste water sample in graduated cylinder .
2- Add 20 ml digestion reagent (H2SO4 +K2SO4 +CuSO4)
3- Place the graduated cylinder in digestion apparatus and leave the sample for digestion
until it reduce to 20-25ml.
4- Add 140ml of distilled water in it
5- Add 20 ml of Hydroxide and Thiosulphate (NaOH+ Na2S2O3)
6- Add phenolphthalein 1ml to give it pink color
7- Take 50 ml of Boric acid (H3PO3) in separate flask.
8- Place the flask and cylinder in Kjeldel apparatus.
9- Leave it until the solution become green color
10- Titrate it with 0.02N H2SO4 till violet color achieve.

6.3 OBSERVATION AND CALCULATION

A= ml of titrant used= 18.6 ml
( A ) x Normaility of titrant x molecular weight of Nitrogen x 1000
Nitrogen(mg/l) =
ml of sample
( A ) x 0.02 x 14 x 1000
Nitrogen (mg/l) =
140

P a g e 28 | 34
 Nitrogen (mg/l) = 37.8 mg/l
The amount of Kjeldel Nitrogen in given water sample is found to be 37.8 mg/l
6.4 COMMENTS:

After performing this experiment we come to know what is kjeldel nitrogen basically
is. And experiment for the determination of kjeldel nitrogen. Concentration of nitrogen in
given water sample is beyond the permissible limits regulated buy WHO so it is not suitable
for drinking it requires treatment before drinking

STUDY OF INSTRUMENTS
8.1 ATOMIC ABSORBANCE SPECTROPHOTOMETER (AAS)
Atomic Absorption Spectroscopy is an instrumental analysis technique for rapid trace
metal analysis. It is based on element specific wavelength light absorption by ground state

P a g e 29 | 34
atoms in the flame or electro thermal graphite furnace. It finds immense applications in the
analysis for trace metals in soils, lakes, rivers, oceans, and drinking water, pharmaceuticals,
foods and beverages, geological and mineralogical samples, petroleum products, biological
fluids and specimens and forensic analysis. It is common to get results in ppm levels and a
higher sensitivity of ppb levels when we using graphite furnace atomization.
8.1.1Atomic Absorption Spectroscopy (AAS)

AAS was employed in the 1950’s Used for qualitative and quantitative detection. It’s
used for the determination of the presence and concentrations of metals in liquid samples.
Metals that can be detected include Fe, Cu, Al, Pb, Ca, Zn, Cd and many more.
Concentrations range is in the low mg/L (ppm) range.

Figure 8.1 Detectable element by ASS

P a g e 30 | 34
 Figure 8.2 Schematic diagram of AAS

1-Light source
The disadvantage of both the HCL and laser is that they have narrow-band light sources and

so only one element is measurable at a time.

1- Hollow cathode lamp

Consists of a cathode and an anode. The cathode is made of the element of interest

1- A large voltage across the anode and cathode will cause the inert gas to ionize.

2- The inert gas ions will then be accelerated into the cathode, sputtering off atoms

from the cathode.

3- Both the inert gas and the sputtered cathode atoms will in turn be excited by

collisions with each other. Lamp (Hollow Cathode Lamb):

4- When these excited atoms decay to lower energy levels they emit a few spectral

lines characteristic of the element of interest.

P a g e 31 | 34
 5- The light is emitted directionally through the lamp's window, a window made of a

glass transparent in the UV and visible wavelengths.

6- The light can then be detected and a spectrum can be determined.

Figure 8.3 Hollow cathode tubes

2- Laser
They are intense enough to excite atoms to higher energy levels. This allows AA and
atomic fluorescence measurements in a single instrument.

Figure 8.4 Laser

2-Atomizer
There are three types of atomizer
1- Flame
2- Hybrid
3- Graphite Furnace

3-Separation of light
Monochromator are used for light separation

P a g e 32 | 34
The light passes from the HCL through the element in the sample to the monochromator. It’s
function is: It isolates the specific light of the element of interest from the other background
lights and transfers it to the photomultiplier tube (detector).

Figure 8.5 Monochromator

4-Detector (photomultiplier tubes):

The PMT detects the amount of reduction of the light intensity due to absorption by the
analyte, and this can be directly related to the amount of the element in the sample. The PMT
converts the light signal into an electrical signal and a computer system translates it into
absorbance. The PMT detects the amount of reduction of the light intensity due to absorption
by the analyte, and this can be directly related to the amount of the element in the sample.
The PMT converts the light signal into an electrical signal and a computer system translates it
into absorbance.

Figure 8.6 PMT

8.2 TOTAL ORGANIC CARBON ANALYZER

Shimadzu has been a world leader in the manufacture of high-quality Total Organic Carbon
(TOC) analyzers since 1967. These state-of-the-art TOC analyzers maximize both sensitivity
and productivity, making them the ideal choice for monitoring rivers, lakes, and oceans,
assisting in the management of water from public sources, wastewater, and manufacturing
P a g e 33 | 34
processes, and, in the pharmaceutical industry, performing validation processes to meet the
required regulations. Today, Shimadzu´s TOC analyzers are the benchmarks of the industry. 

Since all TOC analyzers only actually measure total carbon, TOC analysis always requires
some accounting for the inorganic carbon that is always present. One analysis technique
involves a two-stage process commonly referred to as TC-IC. It measures the amount of
inorganic carbon (IC) evolved from an acidified aliquot of a sample and also the amount of
total carbon (TC) present in the sample. TOC is calculated by subtraction of the IC value
from the TC the sample. Another variant employs acidification of the sample to evolve
carbon dioxide and measuring it as inorganic carbon (IC), then oxidizing and measuring the
remaining non-purgeable organic carbon (NPOC). This is called TIC-NPOC analysis. A more
common method directly measures TOC in the sample by again acidifying the sample to a pH
value of two or less to release the IC gas but in this case to the air not for measurement. The
remaining non-purgeable CO2 gas (NPOC) contained in the liquid aliquot is then oxidized
releasing the gases. These gases are then sent to the detector for measurement.

Whether the analysis of TOC is by TC-IC or NPOC methods, it may be broken into three
main stages:

1. Acidification
2. Oxidation
3. Detection and Quantification

Figure 8.7 TOC

P a g e 34 | 34