DETERMINATION OF pH

Aim: To determine the pH of a given water sample

Principle:
pH is a logarithmic scale and an increase or decrease of one pH unit is a 10-fold change, 

pH = - log10 [H+]

where [H+] = the concentration of Hydrogen ions in moles

The pH can be measured either by the indicator method or by the electrometric method using a pH
meter.
Electrometric method requires a pH meter which is a sensitive instrument to monitor the milli volts
generated by Hydrogen ions and enable the conversion of produced milli volts to corresponding
equivalent pH value in the digital format.
The probe has a sensor bulb of glass containing a fixed concentration of HCl or a buffered chloride
solution in contact with an internal reference electrode. When immersed in solution, the outer bulb
surface becomes hydrated and exchanges sodium ions.  This, together with the repulsion of anions
by fixed, negatively charged silicate sites, produces a potential that is a function of hydrogen ion
activity in solution.

Apparatus:
 Digital pH meter, Beaker, Measuring flask
 Standard solutions for calibration
 Buffer solution of known pH i.e., 4.0, 7.0 & 9.1

Calibration of pH meter:
 The pH meter should be properly assembled and turned on. Usually, pH probe is kept in a
solution of KCl with pH around 4. This KCl will help in maintaining constant potential.
 The instrument should be turned on and the probe should be removed, and cleaned with
distilled water.
 Beforehand, the pH solution containing 4.0,7.0,9.18 has to be made with the help of pH
tablets and stored in storage bottle.
 After turning on the instrument press F2 to calibrate and dip the probe in solution with pH
7.0. After the stable reading comes, press enter and then clean the probe with distilled
water and wipe with tissue.
 Repeat step 4 for pH of 4 and 9.18.
 Then calibration is completed and the pH can be measured.

Procedure:
 After the pH meter is calibrated, the probe has to be cleaned with distilled water and wiped
with tissue. 
  Some amount of unknown sample has to be taken and kept on stirrer so that proper mixing
takes place. 
 Should wait until stable reading appears in the pH meter.
 Three trials have to be done for each sample and readings should be noted down in the
table.
 Every time it should be ensured that probe is cleaned properly.
 After completing the experiment, the probe has to be dipped in KCl solution and the parts
should be removed carefully and placed in their respective positions in the provided box.

Caution: Never leave the probe in dry condition, always dip in buffer solution.

Observation and calculation:

pH meter Reading
S. No. Sample Average
Trial 1 Trial 2 Trial 3

1. Tap water 7.59 7.59 7.59 7.59

Results:
pH of given sample is 7.59.
 Determination of Conductivity using Digital Conductivity Meter

Aim: To analyse the conductivity of given water sample.

Principle:
 Conductivity (k) is a measure of the capability of a solution to pass electric current.
 Varies both with number and types of ions in the solution, which in turn is related to the
concentration of ionized substances in the water.
 Most dissolved inorganic substances in water are in the ionized form and hence contribute
to conductance.
 It is useful for detection of impurities in water.
 Used for quantitative measurement of ionic constituents dissolved in water, which are
important for boiler feed water and cooling water etc.

Apparatus & Reagents:

 Conductivity Meter, Flask, Beaker, Wash Bottle
 Reagents: 0.01 N, KCL solution: Dissolve 0.746 grams of potassium chloride in 1 litre of
distilled water

Procedure:
 First the instrument has to be set properly and then the electrode has to be immersed in KCl
solution and then calibrated. 
 It has to show the reading of 1412 micro mhos at 25°C temperature.
 Now the instrument is calibrated.
 Then the electrode should be removed from KCl solution and cleaned with distilled water
and wiped with tissue.
 Then the electrode has to be dipped in unknown sample and measure the conductivity.
 Three trials should be done.

Observation and calculation:

Conductivity meter Reading
S. No. Sample Average
Trial 1 Trial 2 Trial 3

1. Tap water 1.215 1.215 1.215 1.215

Results:
Electrical Conductivity of given sample is 1.215 milli seimens.
 Hardness Test

Aim: To determine Total hardness, Calcium hardness and Magnesium hardness for a given water
sample.

Principle:
Total hardness may be determined by performing a complexometric titration with EDTA as the
chelating agent. The indicator Eriochrome Black-T (EBT) is normally blue in color but becomes red in
color when it is complexed with calcium or magnesium. Thus, when EBT is added to a solution
containing hardness, it complexes Ca+2 and/or Mg+2 and becomes red in color.
When EDTA, which has much stronger affinity for Ca +2 and Mg+2 than EBT, is added to the solution, it
chelates the Ca+2 and Mg+2 ions complexed with EBT. When all such ions are chelated, i.e., the
endpoint of the titration is reached; EBT reverts to its original blue color.

Equation: EDTA + Ca+2 → Ca – EDTA

Equation: EDTA + Mg+2   →    Mg – EDTA

Apparatus:
 Burette, pipette, measuring cylinders and conical flasks
 Reagents:
1. Ammonia Buffer solution: Dissolve 16.9 g of NH 4Cl in 143 ml concentrated NH4OH.
Add 1.25 g of Magnesium salt of EDTA and dilute to 250 ml with distilled water.
2. Erichrome black T Indicator.
3. Standard EDTA solution 0.01 M: Weigh 3.723 g of AR grade disodium ethylene –
diamine tetraacetate di-hydrate and dissolve in 1 litre of distilled/deionized water.
Check the strength by standardizing with a calcium solution.
4. Standard calcium solution.

Observation and calculation:

Burette Reading
S.No Volume of sample Type of hardness The volume of EDTA consumed
Initial Final Difference

1. 20 ml Total 17 22.7 5.7 5.7 ml

2. 20 ml Calcium 28.4 31.5 3.1 3.1 ml

Blank

Total Hardness
Hardness (in mg/L as CaCO3) = (V × N × 50 × 1000) / (SV) 
Hardness (in mg/L as CaCO3) = (5.7 x 0.01 x 50 x 1000)/20 = 142.5 mg/L
Where: V = volume of titrant (mL); N = normality of EDTA; 50 = equivalent weight of CaCO 3; 
SV = sample volume (mL)

Calcium Hardness
Calcium hardness (in mg/L as CaCO3) = (V × N × 50 × 1000) / (SV) 
Calcium hardness (in mg/L as CaCO3) = (3.1 × 0.01 × 50 × 1000) / 20 = 77.5 mg/L 

Where: V = volume of titrant (mL); N = normality of EDTA; 50 = equivalent weight of CaCO 3; 
SV = sample volume (mL)

Magnesium Hardness
Magnesium Hardness as mg/l as CaCO3 = Total Hardness – Calcium Hardness
Magnesium Hardness as mg/l as CaCO3 = 142.5 – 77.5 = 65 mg/L

Results:
Total hardness of the given water sample as CaCO 3 is = 77.5 mg/L.
 Chlorides Test

Aim: To determine the chloride concentration of a given water sample.

Principle:
Chloride ion concentration in the water can be determined by titration with silver nitrate. As the
silver nitrate solution is slowly added, a precipitate of silver chloride forms.

Ag+ (aq) + Cl– (aq) → AgCl (s)

The end point of the titration occurs when all the chloride ions are precipitated. Then additional
silver ions react with the chromate ions of the indicator, potassium chromate, to form a red-brown
precipitate of silver chromate.

2Ag+ (aq) + CrO42– (aq) → Ag2CrO4 (s)

Titration should be carried out under conditions of pH 6.5 – 9. At higher pH silver ions may be
removed by precipitation with hydroxide ions, and at low pH chromate ions may be removed by an
acid-base reaction to form hydrogen chromate ions or dichromate ions, affecting the accuracy of the
end point.

Apparatus & Reagents:

 Burette, pipette, measuring cylinders and conical flasks
 Reagents:
1. Indicator 5% K2CrO4: 1.0 g of K2CrO4 was dissolved in 20 mL of distilled water.
2. AgNO3 solution: 9.0 g of AgNO3 was weighed out, transferred to a 500 mL volumetric
flask and made up to volume with distilled water. The resulting solution
was approximately 0.1 M.

Procedure:
 Take 100 mL of the sample or take appropriate amount and dilute it to 100 mL .
 Fill burette with silver nitrate titrant.
 Add 2 to 3 drops of the potassium chromate indicator to the sample.
 Titrate the blank with standard AgNO 3. The end point is the change of color from yellow to
brick-red (B).
 Titrate the sample in the same way to the same brick-red color (use blank titration as
reference color and be consistent in end-point recognition) (A).
 Calculate ppm Cl- and record with one decimal.
Observations and calculations:
Burette Readings
S.No. Volume of Sample (ml) Volume of EDTA added (ml)
Initial (ml) Final (ml)

1. 20 12 14.9 2.9
2. 20 14.9 18 3.1

Calculations:
 If Sample is diluted with distilled water,
ppm Cl- = (A-B) x N AgNO3 x 35450 / mL of sample
A = mL AgNO3 consumed for sample.
B = mL AgNO3 consumed for Blank.

 If Sample not diluted with distilled water,

ppm Cl- = mL AgNO3 consumed for sample x N AgNO3 x 35450 / mL of sample
ppm Cl- = (3 x 0.02 x 35450) / 20 = 106.35 mg/L
 
Result:
 The amount of chlorides present in the given sample = 106.35 mg/L.
 
 Alkalinity Test

Aim: To determine the alkalinity of the given water sample.

Principle:
Alkalinity of water can be determined by titrating the water sample with sulphuric acid of
known values of pH, volume and concentration.
When water sample has a pH greater than 4.5 is titrated with acid to end point of pH 4.5, all OH-,
CO32-, HCO3- neutralised.
For the pH more than 8.3, phenolphthalein indicator is used, the colour changes from pink to
colourless. This pink colour is due to presence of hydroxyl ions. If sulphuric acid added to it, pink
colour disappears i.e. OH- ions neutralised.
When methyl orange added to the same sample, the presence of CO32-, HCO3– in the sample changes
the colour to yellow, while adding sulphuric acid the colour changes to orange, this indicates all
CO32-, HCO3- neutralised.

Apparatus & Reagents:

 pH meter
 Burette
 Pipette
 Measuring Cylinders
 Conical Flask
 Reagents:
1. Standard Sulphuric Acid, 0.02N
2. Phenolphthalein Indicator
3. Methyl Orange Indicator
4. Rinse and fill the burette with 0.02N sulphuric acid.
5. Using measuring cylinder take 100ml of sample and pour it in conical flask. (If sample is
highly alkaline dilute the sample)
6. Add 1-2 drops of phenolphthalein indicator to the sample. The colour of the solution
turns pink.
7. Titrate it against 0.02N H2SO4 till pink colour disappears (V1).
8. Add 1-2 drops of methyl orange indicator to the same sample. Titrate it until colour
changes from yellow to orange (V2).

NOTE: If pink colour does not appear after addition of phenolphthalein indicator, continue titration
as explained earlier.
Observation and Calculation:
Volume of 0.02N H2SO4 Consumed (ml)
S.No Volume of Sample
. (ml) Initial Phenolphthalein Point (V1) Methyl Orange Point (V2)

1. 20 11.3 11.3 15.2

2. 20 15.2 15.2 18.9

 Phenolphthalein Alkalinity as CaCO3 in mg/L = (V1×N×50×1000)/Volume of sample

 Total or Methyl orange Alkalinity as CaCO3 in mg/L = (V2×N×50×1000)/Volume of sample

Total or Methyl orange Alkalinity as CaCO3 in mg/L = (3.8 × 0.02 × 50 × 1000)/20 = 190mg/L

Where,
 V1 = mL of standard acid used for phenolphthalein end point
 V2 = mL of standard acid used for methyl orange end point
 N = Normality of standard acid

Summary and type of alkalinity in water

Result of Titration Hydroxide alkalinity as CaCO3 Carbonate Alkalinity Bicarbonate
as CaCO3 alkalinity as CaCO3

P=0 0 0 T
P<½T 0 2P T - 2P

P=½T 0 2P 0
P>½T 2P -T 2(T - P) 0

P=T T 0 0
P = Phenolphthalein Alkalinity, T = Total Alkalinity

Result:
P. Alkalinity as CaCO3 (mg/L) = 0 mg/L
“M.O” or Total Alkalinity as CaCO3 (mg/L) = 190 mg/L
(OH)- Alkalinity as CaCO3 (mg/L) = 0 mg/L
(CO3)2- Alkalinity as CaCO3 (mg/L) = 0 mg/L
(HCO3)- Alkalinity as CaCO3 (mg/L) = 190 mg/L
 Determination of Sulphates

Aim: To measure the concentration of sulphates in the water sample.

Apparatus and regents:

Turbidity meter, pipette, measuring cylinders and conical flasks
Reagents:
 Conditioning agent: Mix 50 ml glycerol with a solution containing 30 ml of concentrated HCl,
300 ml distilled water, 100 ml 95% ethyl or isopropyl alcohol and 15 g NaCl.
 Barium Chloride crystal: 20 to 30 mesh size.
 Standard Sulphate solution: Dissolve 1.479 g of anhydrous Na 2SO4 in 1 litre distilled water.
This suspension contains 1 ml = 100 μg Sulphate.

NOTE: Turbidity meter: Need to be calibrated before use.

Procedure:
 Measure 100 ml water sample or suitable portion of the sample made to 100 ml with
distilled water. 
 Add 5 ml conditioning agent and a spoon full of BaCl2 crystals. Stir exactly for 1 minute. 
 Pour the solution into the glass cell of the turbidity meter and measure the turbidity. 
 Prepare a calibration graph by using sulphate standard as described earlier. 
 The standards can be made from 0 to 40 mg/l sulphate range by taking a 0-40 ml standard
solution and making up to 100 ml. 
 Determine the concentration of a given sample from the calibration chart plotted with
known sulphate concentrations Vs turbidity. 

Result:
Sulphate concentration in given water sample: 4.58 ppm.
 Biochemical Oxygen Demand Test

Aim: Determination of BOD by measuring the DO through titrimetric-based Winkler’s method with
azide modification/membrane electrode method

Apparatus/glassware:
 BOD bottles of capacity 300 mL
 Conical flask of capacity 500 mL
 Burette
 Pipette 
 BOD Incubator
 Reagents:
1. Alkali-iodide-azide reagent
2. Manganese sulphate solution
3. Concentrated Sulphuric acid
4. Sodium thiosulphate solution (0.025N)
5. Starch Indicator.
6. Dilution water (1 Litre containing following reagents)
7. 1 mL of Ferric chloride
8. 1 mL of Calcium chloride
9. 1 mL of Magnesium sulphate
10. 1 mL of Phosphate buffer 
11. Seed- Bacteria from mostly secondary treatment plant or unseeded for the wastewater
which contains bacteria inherently.

Principle:
 2 MnSO4 + O2 → 2 MnO2
 Mn2+ + 2OH- + 1/2O2 → MnO2 + H2O (Brown Precipitate- In presence of O2)
 Mn(OH)2 + 1/2O2 → MnO2 + H2O(Brown Precipitate- In presence of O2)
 Mn2+ + 2OH- → Mn(OH)2 (White Precipitate- In absence of O2)
 - + 2+
MnO2 + 2I + 4H → Mn + I2 + 2H2O (During addition of acid)
 -
I2 + I → I 3 (Iodine will be fixed)
 This iodine is titrated by standard 0.025 N Sodium thiosulfate solution using starch as an
indicator. (1/8 (equivalent wt. of O2) = 0.125 N of Na2S2O3) → 1 ml of sample gives 1 mg of
DO
 N/8 x 1/5 = N/40= 0.025 N Na2S2O3

Azide Modification of Winkler’s method

Problems due to nitrates and iron in sample- it can oxidize I - to I2 in acidic condition leading to over
estimation of BOD,
 NO2- + 2I- + 4H+ → I2 + N2O2 + H2O (Under acid)
 N2O2 + ½ O2 + H2O → 2NO2 + 2H
- +

 NaN3 (azide) + H+ → I2 + HN3 + Na+

  HN3 + NO2- + H+ → N2 + N2O + H2O (Nitrate will be removed)

Experimental Procedure:
 Select required dilutions (to get a minimum final DO of 1.5 mg/L after incubation) and
prepare three BOD bottles to find initial as well as final dissolved oxygen.
 Use bottle one for finding initial DO and keep the other two bottles in the incubator for three
days at 27±1º C or five days at 20±1º C. 
 Determine the DO of first bottle and note as initial DO. Ensure that the water seal is
maintained in the annular space between the stopper and bottle mouth to avoid diffusion of
oxygen from the atmosphere. 
 After 3 days of incubation, find out the DO of incubated samples and note as final DO.
Calculate the BOD of the samples as per the given formula.
 Final DO should be more than 1 - 1.5 mg/L
 At least 2 mg/L of DO should be consumed by organisms after 5 days.

Observation and calculation:

S.No. Type of Sample Volume of Thiosulphate consumed (ml)

Initial Final Difference

1. Wastewater (Day 0) 0 4 4

2. Blank (Day 0) 7.5 11 3.5

3. Wastewater (Day 5) 4.5 8.5 4

4. Blank (Day 5) 11 14.7 3.7

5-day BOD = BOD5 (mg/l) = {[D1-D2]-[B1-B2]}/P

5-day BOD = BOD5 (mg/l) = {[4-3.5] - [4-3.7]}/0.05 = 4mg/L

Where: D1 = DO of the sample immediately after preparation, mg/L,

D2 = DO of the sample after 5-day incubation at 20°C, mg/L, B1 = DO of blank immediately after
preparation, mg/L,
B2 = DO of blank after 5-day incubation at 20°C, mg/L, P= dilution factor

Result:
The BOD of given wastewater sample = 4 mg/L.