LAB MANUAL

ENVIRONMENTAL ENGINEERING

CIVIL ENGINEERING DEPT.

LIST OF EXPERIMENT

1. To Determine biochemical oxygen demand (BOD) of given water/Waste

water sample

2. To determine the chemical oxygen demand (COD) of given sample .

3. To determine the optimum dose of coagulants.

4. Measurement of SO2 in the ambient air.

5. To determination of kjeldahl nitrogen.

6. Determination of alkalinity of given sample of water in mg/l.

7. Determination of chloride content of the given sample.

8. Determination of hardness of the given sample.

9. Determination of PH of given sample.

10. Determination of residual chlorine present in the given sample.

11. Determination of turbidity of the given sample.

12. Determination of most probable number of coli forms.

13. Determination of dissolved oxygen in a sample of water.

14. Determination acidity of water supply. 15. Determination of colour.

EXPERIMENT NO. 01

AIM: - TO DETERMINE BIOCHEMICAL OXYGEN DEMAND (BOD) OF GIVEN

WATER/WASTE WATER SAMPLE.

Introduction:

The biochemical oxygen demand determination is a chemical procedure for determining the

amount of dissolved oxygen needed by aerobic organisms in a water body to break the organic

materials present in the given water sample at certain temperature over a specific period of

time.

BOD of water or polluted water is the amount of oxygen required for the biological

decomposition of dissolved organic matter to occur under standard condition at a standardized

time and temperature. Usually, the time is taken as 5 days and the temperature is 20°C.

The test measures the molecular oxygen utilized during a specified incubation period for the

biochemical degradation of organic material (carbonaceous demand) and the oxygen used to

oxidize inorganic material such as sulfides and ferrous ion. It also may measure the amount of

oxygen used to oxidize reduced forms of nitrogen (nitrogenous demand).

Environmental significance:

BOD is the principle test to give an idea of the biodegradability of any sample and strength of

the waste. Hence the amount of pollution can be easily measured by it. Efficiency of any

treatment plant can be judged by considering influent BOD and the effluent BOD and so also

the organic loading on the unit.

Application of the test to organic waste discharges allows calculation of the effect of the

discharges on the oxygen resources of the receiving water. Data from BOD tests are used for

the development of engineering criteria for the design of wastewater treatment plants.

Ordinary domestic sewage may have a BOD of 200 mg/L. Any effluent to be discharged into

natural bodies of water should have BOD less than 30 mg/L. This is important parameter to

assess the pollution of surface waters and ground waters where contamination occurred due to

disposal of domestic and industrial effluents. Drinking water usually has a BOD of less than 1

mg/L. But, when BOD value reaches 5 mg/L, the water is doubtful in purity. The

determination of BOD is used in studies to measure the self-purification capacity of streams

and serves regulatory authorities as a means of checking on the quality of effluents discharged

to stream waters.

The determination of the BOD of wastes is useful in the design of treatment facilities. It is the

only parameter, to give an idea of the biodegradability of any sample and self-purification

capacity of rivers and streams. The BOD test is among the most important method in sanitary

analysis to determine the polluting power, or strength of sewage, industrial wastes or polluted

water. It serves as a measure of the amount of clean diluting water required for the successful

disposal of sewage by dilution.

Guideline:

According to Bangladesh Environment Conservation Rules (1997), drinking water standard

for biochemical oxygen demand (BOD) is 0.2 mg/L (at 20°C). For wastewater effluent

allowable concentration of BOD varies from 50- 250 mg/L depending on discharge point of

the effluent (e.g., inland water, irrigation land, public sewer etc.)

Principle:

The sample is filled in an airtight bottle and incubated at specific temperature for 5 days. The

dissolved oxygen (DO) content of the sample is determined before and after five days of

incubation at 20°C and the BOD is calculated from the difference between initial and final

DO. The initial DO is determined shortly after the dilution is made; all oxygen uptake

occurring after this measurement is included in the BOD measurement.

Since the oxygen demand of typical waste is sever hundred milligrams per liter, and since the

saturated value of DO for water at 20uC is only 9.1 mg/L, it is usually necessary to dilute the

sample to keep final DO above zero. If during the five days of experiment, the DO drops to

zero, then the test is invalid since more oxygen would have been removed had more been

available.

The five-day BOD of a diluted sample is given by,

BOD5 = [DOi- DOf] × D.F. ………………………….(1)

Here,

Dilution factor (D.F.) =

In some cases, it becomes necessary to seed the dilution water with microorganisms to ensure

that there is an adequate bacterial population to carry out the biodegradation. In such cases,

two sets of BOD bottles must be prepared, one for just the seeded dilution water (called the

"blank") and the other for the mixture of wastewater and dilution wader. The changes in DO

in both are measured. The oxygen demand of waste water (BODw) is then determined from

the following relationship:

BODm× Vm= BODw × Vw + BODd × Vd ………………………(2)

Where, BODm, is the BOD of the mixture of wastewater and dilution water

BODd is the BOD of the dilution water alone;

Vw and Vd are the volumes of wastewater and dilution water respectively in the mixture

and

Vm= Vw+ Vd.

Sample handling and preservation:

Preservation of sample is not practical. Because biological activity will continue after a

sample has been taken, changes may occur during handling and storage.

If Analysis is to be carried out within two hours of collection, cool storage is not necessary. If

analysis cannot be started with in the two hours of sample collection to reduce the change in

sample, keep all samples at 4° C.

Do not allow samples to freeze. Do not open sample bottle before analysis. Begin analysis

within six hours of sample collection.

Apparatus:

1. BOD bottle

2. Beaker (250 ml)

3. Measuring cylinder

4. Dropper

5. Stirrer

Reagents:

1. Manganous sulfate solution

2. Alkaline potassium iodide solution

3. 0.025N sodium thiosulfate

4. Starch solution (indicator)

5. Concentrated sulfuric acid

Procedure:

Fill two BOD bottles with sample (or diluted sample); the bottles should be completely filled.

Determine initial DO (DOi) in one bottle immediately after filling with sample (or diluted

sample). Keep the other bottle in dark at 20°C and after particular days (usually 5-days)

determine DO (DOf) in the sample (or diluted sample). Dissolved oxygen (DO) is determined

according to the following procedure:

1. Add 1 mL of manganous sulfate solution to the BOD bottle by means of pipette, dipping in

end of the pipette just below the surface of the water.

2. Add 1 mL of alkaline potassium iodide solution to the BOD bottle in a similar manner.

3. Insert the stopper and mix by inverting the bottle several times.

4. Allow the "precipitates" to settle halfway and mix again.

5. Again allow the "precipitates" to settle halfway.

6. Add 1 mL of concentrated sulfuric acid. Immediately insert the stopper and mix as before.

7. Allow the solution to stand at least 5 minutes.

8. Withdraw 100 mL of solution into an Erlenmeyer flask and immediately add 0.025N

sodium thiosulfate drop by drop from a burette until the yellow color almost disappears.

9. Add about 1 mL of starch solution and continue the addition of the thiosulfate solution until

the blue color just disappears. Record the ml. of thiosulfate solution used (disregard any

return of the blue color)

Calculation:

Dissolved oxygen, DO (mg/L)

= mL of 0.025N sodium thiosulfate added x Multiplying Factor (M.F.)

Calculate BOD of the sample according to Eq. – 1 or Eq. – 2

DATA SHEET :

Table

Sample No

Source of

Sample

Temperature

of Sample (°C)

BOD (mg/L)

EXPERIMENT NO. 02

AIM: - TO DETERMINE THE CHEMICAL OXYGEN DEMAND (COD) OF GIVEN

SAMPLE.

INTRODUCTION:

The chemical oxygen demand (COD) test allows measurement of oxygen demand of the waste

in terms of the total quantity of oxygen required for oxidation of the waste to carbon dioxide

and water. The test is based on the fact that all organic compounds, with a few exceptions, can

be oxidized by the action of strong oxidizing agents under acid conditions.

Organic matter + Oxidizing agent = CO2 + H2O 12.1

The reaction in Eq.-1 involves conversion of organic matter to carbon dioxide and water

regardless of the biological assimilability of the substance. For example, glucose and lignin

(biologically inert substance) are both oxidized completely by the chemical oxidant. As a result,

COD values are greater than BOD values, especially when biologically resistant organic matter

is present.

Thus one of the chief limitations of COD test is its inability to differentiate between

biodegradable and non-biodegradable organic matter. In addition, it does not provide any

evidence of the rate at which the biologically active material would be stabilized under

conditions that exist in nature.

The major advantage of COD test is the short time required for evaluation. The determination

can be made in about 3 hours rather than the 5-days required for the measurement of BOO. For

this reason, it is used as a substitute for the BOD test in many instances.

Environmental Significance:

"COD is often measured as a rapid indicator of organic pollutant in water; it is normally

measured in both municipal and industrial wastewater treatment plants and gives an indication

of the efficiency of the treatment process. COD has further applications in power plant

operations, chemical manufacturing, commercial laundries, pulp & paper mills, environmental

studies and general education.

Guideline:

According to Bangladesh Environment Conservation Rules (1997), drinking water standard for

chemical oxygen demand (COD) is 4.0 mg/L. For wastewater effluent allowable concentration

of CBOD varies from 200- 400 mg/L depending on discharge point of the effluent (e.g., inland

water, irrigation land, public sewer etc.)

Principle:

Potassium dichromate or potassium permanganate is usually used as the oxidizing agent in the

determination of COD. In this class potassium permanganate would be used in the

determination of COD. Potassium permanganate is selective in the reaction and attacks the

carbonaceous and not the nitrogenous matter.

In any method of measuring COD, an excess of oxidizing agent must be present to ensure that

all organic matter is oxidized as completely as possible within the power of the reagent. This

requires that a reasonable excess be present in all samples. It is necessary, therefore, to measure

the excess in some manner so that the actual amount can be determined. For this purpose, a

solution of a reducing agent (e.g., ammonium oxalate) is usually used.

Apparatus:

1. Beaker (250 mL)

2. Dropper

3. Stirrer

Reagent:

1. Diluted sulfuric acid solution

2. Standard potassium permanganate solution

3. Standard Ammonium Oxalate solution

Procedure:

1. Pipette 100 mL of the sample into a 250 mL Erlenmeyer flask.

2. Add 10 mL of diluted sulfuric acid and 10 mL of standard KMn04 solution.

3. Heat the flask in a boiling water bath for exactly 30 minutes, keeping the water in the bath

above the level of the solution in the flask. The heating enhances the rate of oxidation

reaction in the flask.

4. If the solution becomes faintly colored, it means that most of the potassium permanganate

has been utilized in the oxidation of organic matter. In such a case, repeat the above using a

smaller sample diluted to 100 mL with distilled water.

5. After 30 minutes in the water bath, add 10 mL of standard ammonium oxalate

[(NH4)2C204] solution into the flask. This 10 mL ammonium oxalate, which is a reducing

agent, is just equivalent to the 10 mL potassium permanganate (oxidizing agent) added

earlier. The excess of reducing agent [(NH4)2C204] now remaining in the flask is just

equivalent to the amount of the oxidizing agent (KMn04) used in the oxidation of organic

matter.

6. The quantity of ammonium oxalate remaining in the flask is now determined by titration

with standard potassium permanganate. Titrate the content of the flask while hot with

standard potassium permanganate to the first pink coloration. Record the mL of potassium

permanganate used.

Calculation:

COD (mg/L) =

DATA SHEET

Table

Sample No

Source of

Sample

Temperature of

Sample (°C)

COD (mg/L)

EXPERIMENT NO. 03

AIM: - DETERMINATION OF OPTIMUM DOES OF COAGULANTS

Principle:

The two basic terms which can exactly explain the happenings of this experiment are

“Coagulation” and “Flocculation“.

1. Coagulation: It is the process of addition of a chemical to de-stabilize a stabilized charged

particle.

2. Flocculation: It is a slow mixing technique which promotes agglomeration and helps the

particles to settle down.

* Generally we encounter very fine and charged clay like particles in water treatment which

should be removed before we continue for further processes. These impurities do not settle by

gravity when the water is passed through a sedimentation tank. The reason being that these are

charged particles, they repel each other and just stay.

* The presence of these very fine charged particles increases the turbidity of the water which is

undesirable and hence these impurities are to be removed. Therefore which will be using a

chemical which dissociates as soon as it added to water and helps in the process of

“Coagulation”. In the present experiment we are using “Alum” [Al2(SO4)3. 18H2O] as

the clarifying agent.

*When alum solution is added to water, the molecules dissociate to yield SO4^2- and Al3+.

These charged species combine with the charged colloidal particles to neutralize the charge. A

detailed explanation of the charge removal can be found in the web which will be based on two

basic definitions “Stern potential“ and “Zeta Potential“.

* Through the slow mixing or so called “Flocculation” a process known as agglomeration occurs

which combines the charged particles into a compact whole and helps in the settling of the

particle. That is the reason why we have step of “slow mixing” in the present experiment.

Apparatus:

1. Jar testing apparatus

2. Turbidity meter

3. Beaker, burette, pipette

Reagents required: Alum solution (1 ml containing 10 mg of alum)

Procedure:

1. Take 1000 ml of given sample in 6 beakers.

2. Find the pH of the sample and adjust it to 6 to 8.5.

3. Now add 1 ml, 2 ml, 4 ml, 8 ml, 10 ml, and 12 ml of alum respectively in each one of the

beakers.

4. Now insert the paddle of the jar testing apparatus inside the beakers and start it.

5. Initially maintain a speed such that the paddles rotate at an angular velocity of 100 rpm for a

time of 1 minute.

6. Now adjust the speed such that the paddles rotate at 40 rpm/min for a time of 9 minutes.

7. Now allow the beakers to settle down for 10 minutes.

8. Make an observation as of which of the 6 beakers is most clearer. Also measure the turbidity

of each beaker using a turbidity meter and tabulate your results.

9. Plot a graph “Settled turbidity” Vs “Coagulant dosage”.

Result:

The optimum value of coagulant dosage from the graph should be reported.

* The optimum value of coagulant generally lies between 8 to 10 for normal water from rivers.

The graph varies as shown (only shape not exact graph)

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EXPERIMENT NO.04

AIM: MEASUREMENT OF SO2 IN THE AMBIENT AIR.

Method of measurememt:

Volumetric – sampling a volume of air through a collecting medium at a known flow rate for

a specified time.

Apparatus:

Gas sampler of midget impingers or high volume sampler with a gas kit attachment for SO2

Standard glass bubblers, airflow rotameter for measuring flow rate.

Chemica1s:

0.1 N Sodium tetra-chloromercurate, an absorbing reagent

Samp1img 1ocatiom guide1imes:

Sampling station should be located depending upon the objective of measurement campaign

and be kept at an altitude depending upon the type of study region (roadways, industrial area,

disposal sites, residential tract, etc). Generally it is kept at a height of about 3 to 1o m from

the ground level and sufficiently away from the disturbance or direct obstacle from the source

under consideration.

Samp1img frequency guide1imes:

Sampling is carried out for various purposes. The regular monitoring campaign of national

ambient air quality includes measurement of SO2 typically for 2¢ hours at least twice a week

making about 1o¢ samples a year.

Steps for sampling:

i. Prepare absorbing reagent (sodium tetra-chloromercurate) by dissolving 27.2 g mercuric

chloride and 11.7 g sodium chloride in 1 lit of water

ii. Prepare a sampling train of at least 2 gas bubblers (for average reading) properly washed

with distilled water and air dried.

iii. Place bubblers in the sampling system securely connecting to the manifold. Check the

connections of bubblers with the manifold and the inlet and outlet

iv. Fill the bubblers with an absorbing reagent with an amount sufficient to last for 2¢ hours

(approximately 15 ml for 8 hours sampling to 5o ml for 2¢ hours sampling)

v. To eliminate interferences of trace metals, if any, 1 drop of o.o1% EDTA solution may be

added to the reagent prior to sampling; similarly, effect of oxides of nitrogen may also be

eliminated by adding 1 ml of o.o6% sulphamic acid to the reagent at site

vi. Start the sampler and adjust flow rate to 2 lit/min.

vii. Note the flow rate at the end of the desired sampling period and stop the sampler

viii. Transit the sampling train to environmental laboratory carefully with scientific

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precautions and preserve the sample tubes in controlled environmental conditions.

Laboratory ana1ysis:

Method:

Colorimetric – by estimating absorbance of SO2 from the exposed absorbing reagent at 5¢o nm

using spectrophotometer

Chemica1s:

a. Pararosaniline hydrochloride

b. Acid bleached

c. Formaldehyde (o.2%)

d. Sulphamic acid by dissolving o.8 g in 1oo ml distilled water

e. Standard sulphur dioxide solution by dissolving o.¢ g sodium metabisulphite in 25o ml

distilled water

Steps of ana1ysis:

Calibratiou curve:

i. Prepare a standard solution of SO2 concentrations ranging from o to 25 µg SO2 by taking

definite amounts of std. sulphur dioxide solution in a 25 ml volumetric flasks

ii. Add 1¢ ml of absorbing reagent and 1 ml of pararosaniline hydrochloride to each of the

flasks making a total volume of 2o ml.

iii. Measure absorbance for each flasks by spectrophotometer at 5¢o nm wavelength

iv. Plot graph of absorbance v/s concentration

Absorbance in samples:

i. Transfer samples to a 25 ml flask and develop color as done in calibration curve

ii. Measure absorbance at 5¢o nm

iii. Find out the concentration (µgSO2) corresponding to the measured absorbance from

the calibration curve

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Ca1cu1ations:

i. Average flow rate (if there is a significant difference in initial and final flow rates)

ii. Total volume of air sampled (TVA) in m3

= Avg. flow rate (lit)* 1o-3 (m3/lit) * time (hr) * 6o (min/hr)

iii. µgSO2 / TVA

NAAQ€ im 5g/m3:

Averaging

period

Industrial

areas

Residential, rural &

other areas

Sensitive

areas

2¢ hours 12o 8o 3o

Ammua1 8o 6o 15

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EXPERIMENT NO.05

AIM: DETERMINATION OF KJELDAHL NITROGEN

INTRODUCTION

Nitrogen is one of the five major elements found in organic materials such as protein. The

Kjeldahl method of nitrogen analysis is the worldwide standard for calculating the protein

content in a wide variety of materials ranging from human and animal food, fertilizer, waste

water and fossil fuels.

The Kjeldahl method consists of three steps:

A. Digestion

B. Distillation

C. Titration

A. Digestion of the sample

Digestion is the decomposition of nitrogen in organic samples utilizing a concentrated acid

solution. This is accomplished by boiling a homogeneous sample in concentrated sulfuric

acid. The end result is an ammonium sulfate solution. The general equation for the digestion

of an organic sample is shown below:

Protein + H2SO4 → (NH4)2SO4(aq) + CO2(g) + SO2(g) + H2O(g) (1)

Sulfuric acid has been used alone for the digestion of organic samples. The amount of acid

required is influenced by sample size and relative amount of carbon and hydrogen in the

sample, as well as amount of nitrogen. A very fatty sample consumes more acid. Also, heat

input and digestion length influences the amount of acid loss due to vaporization during the

digestion process. Initially an organic sample usually chars and blackens. The reaction may

at first be very vigorous depending on the matrix and the heat input. With organic

decomposition the digestion mixture gradually clears as CO2 evolves.

The problem with using sulfuric acid alone for digestion is very long digestion times result

with many samples due to the slow rate of organic decomposition. The addition of an

inorganic salt to the digest elevates the boiling point of the H2SO4. The solution

temperature of concentrated sulfuric acid alone is about 330° C. Addition of a salt such as

K2SO4 can elevate the solution temperature of the digestion mixture to 390° C or more,

depending on the ratio of salt to acid.

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This significantly increases the rate of organic decomposition in the digestion mixture,

shortening the length of time required for digestion. There are several precautions to keep in

mind concerning salt addition. First, it is possible to raise the solution temperature of the

digestion mixture too much. If the temperature goes much above 400° C during any phase of

the digestion, volatile nitrogen compounds may be lost to the atmosphere.

Remember that as acid is gradually consumed during the digestion process, for the various

reasons mentioned above, the salt acid ratio of the digest gradually rises. This means that the

hottest solution temperatures are attained at the end of the digestion. Heat input,

consumption of acid by organic material and vaporization, salt/acid ratio, digestion length,

and physical design of the Kjeldahl flask, are all interrelated. Each has an effect on the final

solution temperature. A second precaution is that if the salt/acid ratio is too high, a

considerable amount of material will “salt out” upon cooling of the digest. Concentrated acid

pockets can be contained

Within the cake. These can react violently when concentrated base is added in the

distillation process. A certain amount of salting out can be managed by diluting the digest

with water while it is still somewhat warm, but not too hot.

Several catalysts have been employed by Kjeldahl chemists over the years to increase the

rate of organic breakdown during the acid digestion. Mercuric oxide has been the most

effective and widely used. However, mercury forms a complex with ammonium ions during

digestion. The addition of sodium thiosulfate or sodium sulfide after digestion and before

distillation will break the complex and precipitate mercuric sulfide. This is also important

from a safety point of view, as mercury vapor might escape to the atmosphere during the

distillation process. Because of environmental concerns over the handling and disposal of

mercury, other catalysts are coming more into favor. Many methods employ copper sulfate.

Titanium oxide and copper sulfate in combination have been found to be more effective than

copper sulfate alone. Selenium is frequently used. Commercially prepared mixtures of

potassium sulfate and a catalyst are available from laboratory chemical suppliers.

B. Distillation

Distillation is adding excess base to the acid digestion mixture to convert NH4+ to NH3,

followed by boiling and condensation of the NH3 gas in a receiving solution. This is

accomplished by;

1) Raising the pH of the mixture using sodium hydroxide (NaOH solution). This has the

effect of changing the ammonium (NH4+) ions (which are dissolved in the liquid) to

ammonia (NH3), which is a gas.

(NH4)2SO4(aq) + 2NaOH → Na2SO4(aq) + 2H2O(l) + 2NH3(g) (2)

2) Separating the nitrogen away from the digestion mixture by distilling the ammonia

(converting it to a volatile gas, by raising the temperature to boiling point) and then trapping

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the distilled vapors in a special trapping solution of boric acid (H3BO3). The ammonia is

bound to the boric acid in the form of ammonium borate complex.

H3BO3+ NH3 → NH4+ + H2BO3- (3)

The majority of the NH3 is distilled and trapped in the receiving acid solution within the first

5 or 10 minutes of boiling. But depending on the volume of the digestion mixture and the

method being followed, 15 to 150 ml of condensate should be collected in the receiving flask

to ensure complete recovery of nitrogen. Further extension of the distillation times and

volumes collected simply results in more water being carried over to the receiving solution.

Excess water does not change the titration results. Distillation times and distillate volumes

collected should be standardized for all samples of a given methodology. The rate of

distillation is affected by condenser cooling capacity and cooling water temperature, but

primarily by heat input. Typically the heating elements used for distillation have variable

temperature controllers. A distillation rate of about 7.5 ml/minute is most commonly cited in

accepted methods. Connecting bulbs or expansion chambers between the digestion flask and

the condenser is an important consideration to prevent carryover of the alkaline digestion

mixture into the receiving flask. The slightest bit of contamination of the receiving solution

can cause significant error in the titration step. When very low levels of nitrogen are being

determined, it is advisable to “precondition” the distillation apparatus prior to distillation.

This can be done by distilling a 1:1 mixture of ammonia-free water and 50% NaOH for 5

minutes just before sample distillation to reduce contamination from atmospheric ammonia.

C. Titration:

There are two types of titration: back titration, and direct titration. Both methods indicate the

ammonia present in the distillate with a color change and allow for calculation of unknown

concentrations.

REAGENTS AND APPARATUS

Potassium sulfate, K2SO4+Se tablets (2 for each digestion tube)

Concentrated H2SO4 (in the hood)

35 % (w/v) NaOH for each digestion tube (already prepared).

4.0 % (w/v) Boric acid, H3BO3 (already prepared)

M HCl (already prepared, exact concentration will be given)

Methyl orange (in droppers)

Erlenmeyer flasks

Burette

Digestion tubes

Milk: Do not forget to bring any brand of milk.

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PROCEDURE

1) Sample: 5.0 mL fresh cow’s milk in a digestion tube.

2) Reagents for digestion: to each milk sample and also to an empty digestion tube (blank)

add the followings:

2 tablets of K2SO4 + Se catalyst

10.0 mL of concentrated H2SO4 (98%)

3) Digestion: heat for ca. 30 minutes at 420ºC.

ATTENTION: Do not inhale the gases evolve in reaction 1.

4) Cooling and diluting: let the digestion tubes to cool to 50-60 ºC and add to each add 50

mL of distilled water.

ATTENTION: Let the tubes stand in air to cool, cold water may break the tubes.

5) Distillation:

i. Place the digested samples in digestion tubes to the distilling unit and add 50.0 mL of 35%

(w/v) NaOH.

ii. The sample is distilled until 100 mL of distillate are collected in 25.0 mL of 4.0 % (w/v)

boric acid.

6) Titration: add 2-3 drops indicator to the Erlenmeyer flask and titrate it with 0.1 M HCl.

7) Calculate the amount of protein (% protein) and compare the result with the value give on

the milk.

PRE-LAB STUDIES

Read the introduction from the manual.

1) List name of three steps in Kjeldahl Method.

2) What is the purpose of digestion?

3) Describe the digestion procedure.

4) Write three catalysts that can be used in Kjeldahl Method. Which one will be used in this

experiment?

5) Write down the purpose of distillation and explain each step in the distillation by writing

chemical reaction(s).

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POST-LAB STUDIES

1) What is the aim of using K2SO4 in this experiment?

2) What is back titration? How is it used in Kjeldahl Method?

3) Which reagent will be used to change ammonium ions to ammonia? Write down the

reaction?

4) Explain the role of H3BO3 in this experiment.

DATA SHEET FOR KJELDAHL METHOD

Volume of HCl for replicate 1:

Volume of HCl for replicate 2:

% protein in milk sample:

% protein written on the label:

Concentration of HCl, M:

TA`s Name and Signature:

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EXPERIMENT NO. 06

AIM: TO DETERMINE THE ALKALINITY OF GIVEN SAMPLE OF WATER IN

MG/L

PRINCIPLE

Alkalinity is determined by titrating the sample with a standard solution of a Strong mineral

acid to bicarbonate and carbonic acid equivalence point. Alkalinity is expressed in terms of

CaCO3 equivalent. For samples whose PH is above 8.3,Titration is done in two steps. In the

first step the PH is lowered to 8.3, which is Indicated by phenolphthalein indicator losing the

pink colour and becoming Colourless. In the second phase of titration the PH is lowered to

about 4.5, which is indicated by methyl orange indicator changing colour from yellow to

orange red

APPARATUS REQUIRED

1. Burette

2. Pipette

3. Erlenmeyer flask

REAGENTS

1 .Sulphuric acid 0.02N

2. Sodium thiosulphate 0.1

PROCEDURE

1. Take 20 ml of the given sample in Erlenmeyer flask (v)

2. Add 1 drop of 0.1N sodium thiosulphate solution to remove the free chlorine if Present

3. Add 2 drops of phenolphthalein indicator. The sample turn pink if the PH is above 8.3

4. Run down 0.02N standard sulphuric acid till the solution turn to colourless.

5. Note down the volume of H2SO4added (v1)

6. Add 2 drops of methyl orange indicator the sample turns yellow

7. Repeat titration till the colour of the solution turns to orange

8. Note down the total volume of H2SO4 added (v2)

OBSERVATIONS AND CALCULATIONS

Sl. No. Sample

No.

Volume

of Sample

Initial

burette

reading(ml)

Final

burette

reading(ml)

Volume of

H2So4(ml)

Alkalinity

(mg/l)

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Phenolphthalein alkalinity expressed as mg/l (CaCO3)

P= (V1 X 50 X 1000 X 0.02N)/ vol.of sample used (ml)

Methyl orange alkalinity expressed as mg/l (CaCO3)

M= (V2 X 50 X 1000 X 0.02N)/ vol.of sample used (ml)

Total alkalinity expressed as mg/l (CaCO3)

T= (V3 X 50 X 1000 X 0.02N)/ vol.of sample used (ml)

RESULT

Phenolphthalein alkalinity expressed as mg/l (CaCO3) =

Methyl orange alkalinity expressed as mg/l (CaCO3) =

Total alkalinity expressed as mg/l (CaCO3) =

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EXPERIMENT NO.07

AIM: TO DETERMINE THE CHLORIDE CONTENT OF THE GIVEN SAMPLE BY

MOHR’S METHOD

PRINCIPLE

Chloride ion is determined by Mohr’s method ,titration with standard silver nitrate solution in

which silver chloride is precipitated first. The end of titration is indicated by formation of red

silver chromate from excess AgNO3 and potassium chromate used as an indicator in neutral

to slightly alkaline solution.

AgNO3+Cl- → AgCl+NO3

2AgNO3+ K2CrO4 → Ag2CrO4+2KNO3 (Reddish Brown)

APPARATUS

1. Burette

2. Pipette

3. Erlenmeyer flask

REAGENTS

1. Standard silver Nitrate 0.0141N

2. Sodium Chloride 0.014N

3. Potassium Chromate indicator

PROCEDURE

1. Take 20 ml sample in Erlenmeyer flask

2. Adjust its PH to be between 7.0 and 8.0 either with sulphuric acid or sodium

Hydroxide solution. Otherwise, AgOH is formed at high PH level or CrO4 -2 is

Converted Cr2O7 -2 at low PH level

3. Add 1ml of potassium Chromate to get light yellow colour

4. Titrate with Standard silver Nitrate solution till colour change from yellow to brick red

5. Note the volume of silver Nitrate added (A)

6. For better accuracy, titrate distilled water in the same manner

7. Note the volume of silver Nitrate added for distilled water (B)

OBSERVATIONS AND CALCULATIONS

Chloride (mg/l) = [(A-B) X 35.450 X 0.0141 X 1000 ] /Volume of sample(ml)

Where

A=Volume of Silver Nitrate solution consumed in water sample (ml)

B= Volume of Silver Nitrate solution consumed in distilled water sample (ml)

RESULT

Chloride value of sample =

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EXPERIMENT NO. 08

AIM:-TO DETERMINE THE HARDNESS OF THE GIVEN SAMPLE BY EDTA

TITRIMETRIC METHOD

PRINCIPLE

EDTA and its sodium salt form a compound when added to a solution of certain Metal

cations.If a small amount of dye such as Eriochrome black T is added to an aqueous solution

containing small calcium and magnesium ions at a PH of a 10±0.50 the solution become wine

red. If EDTA is added then Ca and Mg will be complexed .When all these two ions are

completed the solution will turn blue. This is the end point of titration. The higher the PH

sharper the end point, however above PH10, there is a danger of precipitation of calcium

carbonate and magnesium hydroxide .Hence the PH is fixed at 10±0.50.

APPARATUS

1. Burette

2. Pipette

3. Erlenmeyer flask

REAGENTS

EDTA Solution 0.01M

PROCEDURE

1. Take 20 ml well mixed sample in Erlenmeyer flask

2. Add 1 to 2 ml buffer solution so as to bring the PH to 10+0.50 or 10-0.50

3. Add 2 drops Eriochrome black T indicator solution. The solution turns wine red

in colour

4. Titrate against standard EDTA till wine red colour just turns blue. Note down the

Volume (v)

OBSERVATIONS AND CALCULATIONS

Hardness as CaCO3 =(v1xsx1000)/V mg/l

Where v1= ml of titrant used

S=mg of CaCO3 equivalent to 1ml of EDTA solution=1mg CaCO3

V=volume of sample

RESULT

1. Hardness as CaCO3 =

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EXPERIMENT NO.09

AIM:-TO DETERMINE THE PH

OF GIVEN SAMPLE USING PH

PAPER AND

DIGITAL PH METER

PRINCIPLE

PH refers to the hydrogen ion activity. It is expressed as the negative logarithm of the

reciprocal of the hydrogen ion activity in moles per litre. It can be measured by PH paper or

electrometrically by measuring of hydrogen ion by potentiometric measurement using a

standard hydrogen electrode and a reference electrode.

APPARATUS REQUIRED

PH meter along with electrodes

Buffer solution

Thermometer

PH paper

REAGENT

STANDARD BUFFER SOLUTION: preparation of buffer solution :standard solution can be

prepared freshly by dissolving the standard buffer tablets or powders(PH 4 and 7.2)

PROCEDURE

USING PH METERS:

Take the liquid sample which the PH is to be determined in a glass beaker. Note the sample

temperature. Rinse the electrode thoroughly with distilled water and carefully wipe with a

tissue paper.dip the electrode in to the sample solution

USING PH METERS:

Dip the PH paper strip in to the solution. Compare the colour given on the wrapper of the P

H

paper book. Note down the PH of the sample along with temperature.

RESULT

PH

value of sample using PH paper =

PH value of sample using P

H meter =

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EXPERIMENT NO. 10

AIM:-TO DETERMINE THE AMOUNT OF TOTAL RESIDUAL CHLORINE

PRESENT IN THE GIVEN SAMPLE OF CHLORINATED WATER BY STARCH

IODIDE METHOD

PRINCIPLE

Chlorine will liberate free Iodine from Potassium Iodide solution at PH 8.0 or

less. The liberated Iodine is titrated against standard sodium thiosulphate with

Starch as indicator

APPARATUS REQUIRED

1. Burette

2. Pipette

3. Erlenmeyer flask

REAGENTS

1. Concentrated Acetic acid

2. Potassium Iodide

3. Sodium Thiosulphate (0.025N)

4. Starch solution

5. Iodine solution (0.025N)

PROCEDURE

1. Take 25 ml of sample in an Erlenmeyer flask

2. Add 5ml of Acetic acid to bring PH 3.0 to 4.0

3. Add 1gm of potassium iodide and mix thoroughly. Yellow colour is obtained

4. Titrate against standard sodium thiosulphate solution in the burette until a pale

yellow colour is obtained

5. At these stages add 1ml of starch indicator and continue the titration till the

blue colour disappears. Note down the volume (vV1)

RESULT

Turbidity of sample =

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EXPERIMENT NO.11

AIM:-TO DETERMINE THE TURBIDITY OF THE GIVEN SAMPLE USING

NEPHELOMETER IN N.T.U

PRINCIPLE

Turbidity can be measured either by its effects on the transmission of light which is termed as

turbidimetry or its effects on the scattering of light which termed as Nephelometer.

Turbidimetry can be used for sample with moderate turbidity and Nephelometer for sample

with low turbidity.Higher the intensity of scattered lights higher the turbidity.

APPARATUS REQUIRED

Nephelometric turbidimeter Cuvettes it takes the samples for measurements.

REAGENTS

• Solution (1) dissolve 1 g hydrazine sulphate in distilled water and

dilute to 100 ml in volumetric flask

• Solution (2) dissolve 10g hexamine LR grade in distilled water

and dilute to 100ml in volumetric flask

• In 100ml volumetric flask, mix 12.5 ml solution (1) and 12.5 ml

Solution (2) .let them stand for 24 hours at 250 dilute to mark and

mix. The turbidity of the suspension is 1000 NTU

PROCEDURE

CALIBERATION:

• Switch on the instrument and keep it on for some time

• Select appropriate range depending upon the expected turbidity of the sample.

• Set zero of the instrument with turbidity free water using a blank solution and adjust 000

with set zero knob.

• Now in another test tube take standard suspension just prepared as above for 0 – 200 NTU

solution as standard.

• Take its measurements and set the display to the value of the Standard suspension with the

calibrate knob.

MEASUREMENTS:

To determine the turbidity of water sample place the sample in the cuvette and note the

displayed reading .if water has high turbidity it can be suitably diluted and must be shaken

before determination.

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CALCULATION:

Turbidity = A (B + C) / C

A = NTU found in diluted sample

B = volume of dilution water

C = sample of volume taken for dilution

RESULT

Turbidity of sample =

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EXPERIMENT NO. 12

AIM: - DETERMINATION OF MOST PROBABLE NUMBER OF COLI FORM

Introduction:

A variety of different microorganisms are found in untreated water. Most of these organisms

do not pose a health hazard to humans. Certain organisms, referred to as pathogens, cause

disease to humans which include species of bacteria, viruses and protozoa. These organisms

are not native to aquatic systems and usually require an animal host for growth and

reproduction. Pathogens are likely to gain entrance sporadically, and they do not survive for

very long period of time; consequently they could be missed in a sample submitted to the

laboratory. Although it is possible to detect the presence of various pathogens in water, the

isolation and identification of many of these is often extremely complicated, time-consuming

and expensive proposition. Hence in most cases (except when presence of any particular

microorganism is suspected) the microbiological quality of water is checked using some

indicator organisms.

An indicator organism is one whose presence presumes that contamination has occurred and

suggests the nature and extent of the contaminants. An indicator organism should be a

microorganism whose presence is evidence of fecal contamination of warm blooded animals.

Indicators may be accompanied by pathogens, but typically do not cause disease themselves.

The ideal indicator organism should have the following characteristics:

1. Always be present when pathogens are present

2. Always be absent where pathogens arc absent

3. Numbers should correlate the degree of pollution

4. Be present in greater number than pathogens

5. There should be no after-growth or re-growth in water

6. There should be greater or equal survival time than pathogens

7. Be easily and quickly detected by simple laboratory tests

8. Should have constant biochemical and identifying characteristics

9. Harmless to humans

No organisms or group of organisms meet all of these criteria; but the coliform bacteria fulfill

most of them, and this group is most common indicator used in microbial examination of

water. 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 are 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

22 | P a g

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higher selective temperature range of 44° to 44.5°C. In addition, they form in hole 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 Thus, the

total coliform group should not be regarded as an indicator of organisms exclusively of fecal

origin. The use of total coliforms as an indicator may therefore be of little value in assessing

the fecal contamination of surface water, unprotected shallow wells etc. where contamination

by coliforms of non-fecal origin can occur. 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 coli (E. coil), which is exclusively of fecal origin, but strains of

Klebsella pneumonia and Enterobacterspecies may also be present in contaminated water.

Using coliform as indicators of the presence and absence of pathogens sometimes may cause

the following drawbacks:

1. False positive result can be obtained from the bacterial genus aeromonas, which can

biochemically mimic the coliform group

2. False negative result can be obtained when conforms are present along with high

population of other bacteria. The latter bacteria can act to suppress coliform activity.

3. A number of pathogens have been shown to survive longer in natural waters and / or

through various treatment processes than coliform

But the use of coliforms was established first and there does not appear to be any distinct

advantages to warrant shifting to other indicator organisms. Since bacteria are used as

indicator organisms, the microbiological examination of water is commonly called

bacteriological examination.

Apparatus:

1. Petri Dish

2. Incubator

3. Measuring Cylinder, beaker, dropper etc.

Reagents:

= Appropriate culture medium (broth)

= Distilled water

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Methods of Bacteriological Examination of Water:

Basically there two methods of bacteriological analysis of water: (a) Multiple Tube or

Most Probable Number (MPN) method, and (b) Membrane Filter (MF) method.

(a) Multiple Tube/ Most Probable Number (MPN) method:

MPN is a procedure to estimate the population density of viable microorganisms in a test

sample. It’s based upon the application of the theory of probability to the numbers of

observed positive growth responses to a standard dilution series of sample inoculums placed

into a set number of culture media tubes. Positive growth response after incubation may be

indicated by such observations as gas production in fermentation tubes or visible turbidity in

broth tubes, depending upon the type of media employed.

(b) Membrane Filter Method: In contrast to the multiple-tube (MT) method, the membrane

filter (IVIF) method gives a direct count of total coliforms and fecal coliforms present in a

given sample of water. The method is based on the filtration of a known volume of water

through a membrane filter consisting of a cellulose compound with a uniform pore diameter

of 0.45 µm; the bacteria are retained on the surface of the membrane filter. When the

membrane containing the bacteria is incubated in a sterile container at an appropriate

temperature with a selective differential culture medium, characteristic colonies of coliforms

and fecal coliforms develop, which can be counted directly. This technique is popular with

environmental engineers. This method is not suitable for turbid waters, but otherwise it has

several advantages. Its particular advantages and limitations are as follow

Advantages:

1. Results are obtained more quickly as the number of coliforms can be assessed in less

than 24 hours, whereas the multiple tube technique requires 48 hours both for a negative

or a presumptive positive test;

2. Saving in work, certain supplies and glassware;

3. Method gives direct results;

4. Easy to use in laboratories, or even in the field if portable equipment is used.

Disadvantages:

1. High turbidity caused by clay, algae, etc. prevents the filtration of a sufficient volume of

water for analysis and it may also produce a deposit on the membrane which could

interfere with bacterial growth;

2. Presence of a relatively high non-coliform count may interfere with the determination of

coliforms:

3. Waters containing particular toxic substances which may be absorbed by the membranes

can affect the growth of the coliforms.

Test Procedure (For MF method):

24 | P a g

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This section describes the general procedures, it should be noted that different types of

filtration units and equipment are available in the market for performing the tests.

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.

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.

25 | P a g

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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:

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:

Calculation:

Total coliform (CFU/ 100 mL) =

Fecal coliform (CFU/ 100 mL) =

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EXPERIMENT 13

OBJECTIVE: - TO DETERMINE DISSOLVED OXYGEN IN A SAMPLE OF WATER.

Apparatus required: Burette, pipette, conical flask, beaker, measuring cylinder

Chemicals required: MnSO4, KOH, KI, Na2S2O3, Starch, and NaN3

Theory:

It is based on oxidation of potassium iodide. The liberated iodine is titrated against standard

hypo solution using starch as a final indicator. Since oxygen in water is in molecular state and

not capable to react with KI, an oxygen carrier manganese hydroxide is used to bring about the

reaction between KI and O2.Manganous hydroxide is produced by the action of potassium

hydroxide and manganous sulphate.

Chemical reaction:

2KOH+MnSO4 = Mn (OH) 2+K2SO4

2Mn (OH) 2+O2 =

2MnO (OH) 2

Mn O(OH)2+H2SO4 =

MnSO4+2H2O+[O]

2KI+H2SO4+ [O] =

K2SO4+H2O+I2

I2+2Na2S2O3=

2NaI+Na2S4O6

Sodium tetrathionate

Starch + I2 Starch iodide complex (Blue in colour)

Procedure:

1. Take 500ml of water in a D.O bottle.

2. Add 10ml of alkaline KI and 10 ml of MnSO4 into it.

3. Stopper the bottle and shake it well.

4. Keep the bottle in dark for 5 min and add conc H2SO4 till the brown precipitates are

dissolved.

5. Take 100 ml of the above solution in a conical flask. Titrate against hypo till the color

changes to light Yellow.

6. Add 3-4 drops of starch in to it and the color changes to blue.

7. The blue color solution is titrated against hypo solution till blue color disappear

8. Stopper the bottle and shake it well.

9. Keep the bottle in dark for 5 min and add conc. H2SO4 till the brown precipitates are

dissolved.

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10. Take 100 ml of the above solution in a conical flask. Titrate against hypo till the color

changes to light Yellow.

11. Add 3-4 drops of starch in to it and the color changes to blue.

12. The blue color solution is titrated against hypo solution till blue color disappeared.

13. This is end point of the titration. Repeat this process till to get three concordant reading.

Tabulation

No. of

Observation

Volume of

water

IBR in ml FBR in ml Diff in ml Remark

Calculation:

100 ml of 1 N Na2S2O3 = 8 gm of O2

V ml of N/40 Na2S2O3 = 8𝑉

40 𝑥 1000 gm of O2 =

8𝑉 𝑥 1000

40 𝑥 1000 mg of O2 =

𝑣

5 mg of O2

100 ml of water sample contain V/5 mg of O2

1 lit of water sample contains = 2V mg of O2 = ppm

Conclusion:

The amount of dissolved oxygen in a sample of water is found to be….. ppm.

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EXPERIMENT 14

AIM: - TO DETERMINE THE ACIDITY OF WATER SUPPLY APPARATUS

REQUIRED:

1. Burette

2. Conical flask

3. Funnel

CHEMICALS REQUIRED:

1. Standard NaOH (N/50)

2. Phenolphthalein

3. Sodium thiosulphate (Na2S2O3)

4. Tap water.

THEORY:

Measurement of acidity is important as acidic water are corrosive and corrosion producing

substances have to be controlled or removed. If the acidic waters are used for stream

generation in boilers, it results in corrosion & decreased efficiency of boiler. For mineral

acids, the titration is carried to a pH

of about 4.5 by using methyl orange indicator, giving a

colour change from red to yellow. The acidity thus determined is called methyl orange

acidity. Total acidity or phenolphthalein in acidity is determined by carrying the titration to

phenolphthalein end point of 8.3 (measuring mineral acids, organic acids and free CO2). The

results are expressed as part of equivalent CaCo3 per million parts of water.

END POINT:

1. Appearance of pink colour with NaOH in the burrete and a drop of phenolphthalein.

2. Change of yellow colour to pink with methyl orange as a indicator

PROCEDURE:

1. Pipette out 100ml of the water sample into a conical flask

2. Add 1 drop of N/10 Na2S2O2 solution to destroy any residual chlorine

3. Add 2-3 drops of methyl orange indicator and titrate against N/50 NaOH solution until

the red colour changes to yellow.

4. Take at least 3 concordant readings and records the volume of NaOH (Sodium

hydroxide) used as Y m

29 | P a g

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TABULATION:

No. of

Vol.of water IBR in ml FBR in ml Diff in ml Remark

Obs

ml

CALCULATION:

Volume of water sample taken for each titration = 100

Volume of N/50 NaOH used in presence of methyl orange indicator = Y ml

Volume of N/10 NaOH used in presence of Phenolphthalein indicator = Z ml

PHENOLPHTHALEIN ACIDITY:

N1V1 = N2V2

Sample = N/50 NaOH

N= Y/50 X 100

Phenolphthalein acid =

PHENOLPHTHALEIN ACIDITY:

1. Proceed as above 2-3 drops of Phenolphthalein indicator (in place of methyl orange) and

titrate the sample against N/50 NaOH (from burette) until the solution is turned to pink

colour and the colour persists for at least 30 second.

2. Repeat the titration to get at least two concordant readings. 3. Record the volume of NaOH used as Z ml. OBSERVATION:

In this titration, the colour changes from colourless to light pink.

CONCLUSION:

The acidity of the given water sample is found to be……………… ppm.

PRECAUTION:

1. The cleaned apparatus should be used for the titration. 2. To avoid loss of CO2, the titration should be carried out quickly & vigorous shaking

should be avoided.

EXPERIMENT NO. 15

AIM: TO DETERMINATION OF COLOUR

INTRODUCTION:

Pure water should not pose any color. Color in water may result from the presence of natural metallic

ions (iron and manganese), humus and peat materials, plankton, weeds, and industrial wastes.

Impurities in water may exist either in the colloidal from or in suspended state. Color caused by

dissolved and colloidal substances is referred as "true color" and that caused by suspended matter, in

addition to dissolved and colloidal matters, is called "apparent color" as it can be easily removed by

filtration. Ground water may show color due to the presence of iron compounds. The color value of

water is extremely pH-dependent and invariably increases as the pH of the water is raised. For this

reason recording pH along with color is advised.

Environmental significance:

Though presence of color in water is not always harmful to human but in most cases it is. Even if the

water is not harmful, aesthetically people do not prefer to use water with color. Moreover,

disinfection by chlorination of water containing natural organics (which produces color) results in

the formation of tri-halomethanes including chloroform and a range of other chlorinated organics

leading to problems which is a major concern in water treatment. So it is important to limit the color

of water for domestic supplies.

Guideline:

According to Bangladesh Environment Conservation Rules (1997), drinking water guideline value

for color is 15 Pt-Co Unit.

Theory on experimental method:

Available methods for determining color of water:

1. Standard Color Solutions Method

2. Dilution Multiple Method

3. Spectrophotometric method

1. Standard color solution method

Waters containing natural color are yellow-brownish in appearance. Standard Color Solution:

Solutions of potassium chloroplatinate (K2PtCl6) tinted with small amounts of cobalt chloride yield

colors that are very much like the natural colors. In this method, the color produced by 1 mg/l of

platinum (as K2PtCl6) and 0.5mg/l of cobalt (as CoCl2•6H2O) is taken as the standard one unit of

color. Usually, a stock solution stock solution of K2PtCl6 that contains 500mg/l of platinum is

prepared, which has a color of 500 units. Then, a series of working standards may be prepared from

it by dilution. Color -comparison tubes are usually used to contain the standards.

31 | P a g

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A series ranging from 0 to 70 color units is employed and samples with color less than 70 units are

tested by direct comparison with the prepared standards. For samples with a color greater than 70

units, a dilution is made with distilled water distilled water to bring the resulting color within the

range of the standards. In this case, the final result should be corrected using a dilution factor.

2. Dilution multiple method

Color of most domestic and industrial waste waters are not yellow-brownish hue.

Other systems of measurement have to be used to measure and describe colors that do not fall into

this classification. For dilution multiple methods, color is measured by successive dilutions of the

sample with color -free water until the color is no longer detectable comparing with distilled water.

The total dilution multiple is calculated and used to express the color degree.

3. Spectrophotometric method

The platinum-cobalt method is useful for measuring color of potable water and of water in which

color is due to naturally occurring materials. It is not applicable to most highly colored industrial

wastewaters. In the laboratory color of water is usually measured using spectrophotometer which

uses light intensity of a specific wavelength (455 nm). The color test measures (inversely) an optical

property of water sample which result from the absorption of light of specific wavelength by the

soluble color substances present in water, Before measuring the color of water it is necessary to plot

standard calibration curve for color using different standard platinum-cobalt solutions of known

concentrations within the range of interest.

Figure 2.1: Optical property used in measurement of color in water using spectrophotometer

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Apparatus:

1. Color Disk

2. Filtration system including filter paper, funnel, holder, beaker etc.

Reagent:

1. Standard potassium chloroplatinate solution

Procedure:

1. Prepare standard samples having color within a specific range by mixing different concentration

of standard potassium chloroplatinate solution with distilled water. Using these samples to

prepare a color calibration curve (absorbance vs. color concentration) for the spectrophotometer.

2. Take 50-mL of filtered test sample in a beaker. Take 50-mL distilled water in another beaker.

Use this sample as blank.

3. Set the spectrophotometer to determine color concentration of the sample.

4. Put the blank sample inside the spectrophotometer cell and set the reading "zero''.

5. Bring out the blank sample and place the test sample inside the spectrophotometer

6. After a While the display will show the color concentration of the sample.

Observation table:

Results:-