Stamford University Bangladesh

Department of Environmental Science

Program: B. Sc in Environmental Science
Course Title: ENV-331b Laboratory Work (Environmental Chemistry)
Course Teacher: Dr. Mahmuda Parvin

Name of the Experiment: Determination of Biochemical oxygen Demand (BOD)

Introduction
When bio-chemical organic matter is discharged into a water body, micro-organisms especially
bacteria, fed on waste, breaking it down into simpler organic and inorganic substances. When
this decompositions takes place in an aerobic environment i.e, in presence of oxygen the process
is “Arobic Decomposition” and produce non-objectionable stable products such as CO 2, SO4,
PO4, NO3.
A simplified representation of aerobic decomposition is given by the following equation

micro-organisms
Organic matter + O2 CO2 +H2O + New cells + Stable Products (1)

When insufficient O2 is available or O2 is exhausted by the aerobic decomposition of wastes,

different sets of microorganisms carry out the decomposition anaerobically (Eq-2) producing
highly objectionable products including H2S, NH4 and CH4.
Organic matter = CO2 + CH4 + New cells + Unstable Products (2)
Considering simpler carbohydrates e.g., C(H2O), as organic matter the (Eq-1) can be written to
represent the process of respiration
The amount of oxygen required by micro-organisms to oxidize organic wastes aerobically is
called biochemical oxygen demand (BOD). BOD is a measure of the strength of a water or
wastewater, the greater the concentration of degradable organic carbon, the higher the BOD.
Domestic and industrial wastes contains high levels of BOD, which if discharged untreated
would seriously deplete DO reserves and reduce the diversity of aquatic life [3].
Theory
An aliquot of water sample is maintained in an incubator at 20c for five days in a closed BOD
bottle, without allowing air to water. During this time the water sample is assumed to be
stabilized, i.e., the bacterial decomposition gets completed. Measuring the dissolved oxygen in
the water sample before and after incubation would indicate the amount of oxygen used for
stabilizing the water.
A water sample with low BOD can be straightaway used for the BOD determination. But a water
sample with a high BOD must be diluted accurately and pretreated before the determination of
BOD [1].
Five-Day BOD Test
The 5-day BOD or BOD5 is the total amount of O2 consumed by the micro-organisms during the
first 5 days of biodegradation. In this simplest form, a BOD5 test would involve putting a sample
of water/ waste water into a stoppered bottle, measuring the dissolved oxygen (DO) of the
sample at the beginning of the test and again at the end of five days. Light must be kept out of
the bottle to keep algae from adding O 2 by photosynthesis and the stopper is used to keep air
from replenishing DO from outside. To standardize the procedure, the test is run at a fixed
temperature of 20C. Since the O2 demand of typical waste is sever hundred milligram per liter,
and since the saturated value of DO for water at 20C is only 9.1 mg/L, it is usually necessary to
dilute the sample to keep final DO above zero. Unit of BOD is mg/L or g/m3
The five-day BOD of a dilute sample is given by
BOD5 = [ DOi – DOf ]  D.F.
Where D.F. = Dilution Factor = (Vol of waste water + dilution water) / (Vol. of wastewater)
Theory of DO Determination
The reactions involves in the various steps of the Winkler Method dissolved oxygen (DO)
determination are presented below:
Manganous sulfate reacts with potassium hydroxide in the alkaline potassium iodide solution to
produce a white precipitate of manganous hydroxide.
MnSO4 + 2KOH = Mn(OH)2 + K2SO4
If the white precipitate is obtained, there was no dissolved oxygen in the sample and there is no
need to proceed further. A brown precipitate is manganic basic oxide and formed as follows:
2Mn(OH)2 + O2 = 2MnO(OH)2
Upon the addition of (sulphuric) acid, this precipitate is dissolved, forming manganic sulfate:
2MnO(OH)2 + 2H2SO4 = Mn(SO4)2 + 3H2O
This compound immediately reacts with potassium iodide, liberating iodine and resulting in the
typical iodine (blue) coloration of the water.
Mn(SO4) + 2KI = MnSO4 + K2SO4 + I2
The quantity of iodine liberated by these reactions is equivalent to the quantity of oxygen present
in the sample. The quantity of iodine is determined by a titrating a portion of the solution with a
standard solution of sodium thiosulfate solution.
2Na2S2O3 + I2 = Na2S4O6 + 2NaI [3]
Required Chemical
 0.5M Manganous sulfate solution
 0.5M Alkaline potassium iodide solution
 0.025N Sodium thiosulfate
 Starch solution (indicator)
 Concentrated Sulfuric acid
Required Instrument:
BOD bottle : 3 nos.
Beaker (250 ml) : 1 no.
Measuring cylinder : 1 no.
Dropper : 1 no.
Stirrer : 1 no.
Procedure
Fill two BOD bottles with samples (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 time (usually 5 days)
determine DO (DOf) in the sample. Dissolved oxygen (DO) is determined according to the
following procedure:
1. Add 1 ml of manganous sulfate (0.5M) solution to the BOD bottle by means of pipette
dipping an end of the pipette just below the surface of water.
2. Add 1 ml of alkaline potassium iodide (0.5M) 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 for at least 5 minutes.
8. Withdraw 100 ml of solution into an Erlenmeyer flask and immediately add 0.025N sodium
thiosulfate dropwise from a burette until the yellow almost disappear.
9. Add about 1ml of starch solution and continue the addition of thiosulfate solution until the
blue color just disappears. Record the ml of thiosulfate solution used [2] [3].

Calculation
Dissolved oxygen, DO (mg/l)  ml of 0.025N sodium thiosulfate added  2
Then BOD can be calculated by the following equation:
BOD5  [DOi  DOf]  DF
Where DF  Dilution factor  (Vol. of wastewater + dilution water)/ (Vol of wastewater) [3]
Or
DOi - DOf
BOD5 
P

P is decimal volumetric fraction of wastewater used in test.’

Volume of sample
P=
Total volume of BOD bottle

References
1. R Gopalan, A Anand and R W Sugumar, Laboratory Manual for Environmental Chemistry,
International Publishers House Pvt. Ltd., New Delhi, pp. 32-34, 2008.
2. The Laboratory Manuals, Department of Applied Chemistry and Chemical Engineering,
University of Dhaka.
3. The Laboratory Manuals of Environmental Engineering, Dr. M. Ashraf Ali Department of
Civil Engineering, BUET, pp. 26-31.