Progressive Education Society’s

MODERN COLLEGE OF ARTS, SCIENCE AND COMMERCE

DEPARTMENT OF BIOTECHNOLOGY
(A DBT-BUILDER, DST-FIST Sponsored Department; supported under
DBT-Star College Scheme)

Aim: To determine biochemical oxygen demand (BOD) given water/wastewater 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 a 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 conditions 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 ions. 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 influential 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 an 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 the 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.

Principle:
The sample is filled in an airtight bottle and incubated at a 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 seven 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.)

Apparatus:
1. BOD bottle
2. Beaker (250 ml)
3. Measuring cylinder
4. Dropper
5. Stirrer

Reagents:
1. Manganous sulfate solution – 1M
2. Alkaline potassium iodide solution – 1M
3. sodium thiosulfate – 0.0125N
4. Starch solution (indicator) (1%– 0.5g in 50 ml starch)
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 2 mL of manganous sulfate solution to the BOD bottle by means of pipette, dipping in
the end of the pipette just below the surface of the water.

2. Add 2 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 2 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 drops by drop from a burette until the yellow colour almost disappears.

9. Add about add few drops of starch solution and continue the addition of the thiosulfate
solution until the blue colour just disappears. Record the ml. of thiosulfate solution used
(disregard any return of the blue colour)

Calculations:
Assumptions for calculations1:
1g equivalent of Sodium Thiosulphate corresponds to 1g of oxygen.
1g equivalent of Na2S2O3 ≡ 1g equivalent of O2
1N Na2S2O3 ≡ 8 g of O2
∴ x ml of ‘y’ N Na2S2O3 ≡ x × y × 8g
1000
Where y = 0.0125 N.

For z ml of sample: x×y×8g×z This value provides us with amount of DO in terms

For obtaining the value of DO in terms of mg/ml multiply the above equation with 1000.

1) For sample 1 (Mula Mutha):

2) For sample 2 (Pawna Lake):

Observation Table:
 Day and DO Distilled water Sample1 (Mula Mutha river) Sample 2 (Pawna lake)
Value (ml) (ml) (ml)
Day1 8.3 ml 1.2 ml 2 ml
DO1 83 mg/ml 12 mg/ml 20mg/ml
Day5 13 ml 0.8 ml 0.5 ml
DO5 130 mg/ml 8 mg/ml 5mg/ml
BOD 4mg/ml 15mg/ml

Result : BOD for sample 1 = 4mg/ml

Interpretation:
According to CPCB (Central Pollution Control Board) water sources must not exceed BOD
level of 3mg/ml. Since both the samples are exceeding the BOD level of 3mg/ml, it signifies
that both the water samples are polluted and require water treatment. Sample from Pawna
lake being highly polluted (BOD = 15mg/ml) in comparison to Mildly polluted sample of
Mula Mutha river (BOD = 4mg/ml) which is being conventionally treated since 2013.