BOD / CBOD

FROM
A TO Z

Amy Starkey
Stark County Sanitary Engineers

What
is
BOD????

What is BOD?
It is a measure of the amount of oxygen consumed
by bacteria during the decomposition of organic
materials.
Organic materials from the wastewater treatment facility
act as a food source for the bacteria.

Directly related to Dissolved Oxygen

The bacteria require oxygen in the form of dissolved
oxygen to decompose or eat the food source. Through a
calculation, the amount of DO depletion between the initial
day and final day of the analysis determines the BOD.
Thus, BOD directly affects the amount of Dissolved
Oxygen
The greater the BOD = more rapid oxygen depletion = less
oxygen available to aquatic life.

What is the Difference Between

BOD and CBOD
BOD
represents the oxidation
of carbons and
nitrogenous compounds
present in the water

CBOD
measures the oxidation of
carbons present in water

TCMP

2-chloro-6-(trichloromethyl)pyridine
prevents the oxidation of reduced forms of nitrogen such
as ammonia, and organic nitrogen which exert a
nitrogenous demand.
Should add at the beginning of the test because
nitrification will begin almost immediately if the right
organisms are present (Baird and Smith, 2002).

BOD
verses
COD

BOD verses COD

BOD
represents the oxidation
of carbons and
nitrogenous compounds
present in the water
Analysis completion is
done in 5-days

COD
Is the measure of the
total amount of oxygen
required to oxidize all
organic material into
carbon dioxide and water
analysis only takes a few
hours

BOD verses COD

COD results are always higher than BOD
results.
Useful in determining an unknown BOD range
for a sample but it can NOT replace the BOD
test.

Methods

Approved Methods
Standard Methods 18th, 19th, and 20th
editions (5210B, 5-Day BOD Test)

Two Ways in Determining DO

Iodometric Method (Winkler DO Method)
Membrane Electrode Method

Winkler Method
Azide Modification Method
Preferred for most wastewaters
Removes interferences caused by nitrite which is common in
wastewaters.

Permanganate Modification
Used when ferrous iron is present

Azide plus Potassium Fluoride Modification

Used when 5 mg or more of ferric iron salts/L are present

Alum Flocculation Modification

Used when there is interference caused by suspended solids

Copper Sulfate-Sulfamic Acid Flocculation

Modification
Used for biological flocs such as activated sludge mixtures which
also have a high oxygen utilization rate

Sample
Collection,
Holding Time,
and
Storage

Grab Samples
Ideally samples should be analyzed within
6-hrs of collection, however if this is not
possible, then analyze samples within 48
hours of collection (40 CFR part 136).
Store samples at < 6C.

Composite Sampling
Samples should be kept at or below < 6C during
compositing (limited to 24-hour period).
start the measurement of holding time from the end of
the compositing period.
For example if the compositing was started at 8:30 am
on Tuesday and ended at 8:30 am on Wednesday, then
the 48-hour holding time would start from the end of the
compositing period which would be 8:30am on
Wednesday.

Store samples at < 6C

Quality
Control &
Procedure
Requirements

BOD Quality Controls

Blank Control Checks
GGA Control Checks
Glucose Glutamic Acid

Seed Control Checks

Blank Control Checks

Straight dilution water
Used to determine cleanliness of bottles
as well as the source water.

It must have a DO uptake NO greater

than 0.2 mg/L

GGA Control Checks

Used to check the quality of the seeding
material.
Low results reflect poor seeding material.

The ideal GGA range for a BOD sample is

198 + 30 mg/L.

GGA Control Checks

GGA Needs to be pH adjusted
Initially the pH is around 4
Adjust between 6.5 -7.5, like any other
samples

Seed Controls
Must have a DO uptake attributable to the
seed added to each bottle between
0.6mg/L 1.0 mg/L.
Most domestic wastewater, unchlorinated
or undisinfected effluents will contain a
sufficient population of microorganisms.
Used to calculate the BOD results of
samples which are seeded

BOD / CBOD Requirements

pH of samples should be between a pH of 6.5-7.5
Sample temperature should be adjusted to 20 + 1C before making
dilutions
After 5 days of incubation the final DO of samples must result in a
DO depletion of at least 2.0 mg/L with a DO residual of no less than
1.0 mg/L. This is why it is recommended to make several dilutions
of a sample.
Example: The initial DO of a sample is 8.2 and after 5 days of
incubation the final DO is 7.8. Then the final DO does not meet the
required DO depletion of at least 2.0mg/L so a BOD result can not be
calculated from this sample.
Example #2: The initial DO of a sample is 8.2 and after 5 days of
incubation the final DO is 0.20 mg/L. Here the final DO does not meet
the required DO residual of at least 1.0 mg/L, so again the BOD result
can not be calculated

Dilution
Water

Dilution Water Sources

Distilled
Tap
De-mineralized
Natural Waters

Dilution Water Source

Must be free of heavy metals, and toxic
substances which inhibit micro-bacterial
growth.
Must also be able to maintain no more
than a 0.20 mg/L DO depletion during the
5-day incubation period.

Reagents Added to Dilution Water

Phosphate Buffer Solution
Magnesium Sulfate Solution (MgSO4)
Calcium Chloride Solution (CaCl2)
Ferric Chloride Solution (FeCl3)

Purpose of Adding Trace Metals,

Nutrients and Buffering Dilution
Water
Bacteria growth requires nutrients and
trace metals.

It is buffered to ensure the pH of the

incubated samples remain in a range
suitable for bacteria growth.

Why Dilute Samples Before

Incubation?
Because the BOD concentration in most
wastewaters exceeds the concentration of
DO available in an air-saturated sample.

Seeding

Why Seed?
To add a population of microorganisms capable
of oxidizing the biodegradable organic matter.
Most domestic wastewater, unchlorinated or
undisinfected effluents will contain a sufficient
population of microorganisms.

Samples That may Require Seeding

Chlorinated samples
High temperature wastes
Wastes with extreme pH values

Selecting a Seed Source

Select a material to be used for seeding which
will have a BOD of at least 180 mg/L.

Example of seed sources according to Standard

Methods 20th Edition
Raw domestic Sewage prepared as stated above
Small quantities of digester supernatant, return
activated sludge
Commerically available seed material (Polyseed)

Seeding
Must have a DO uptake/depletion of 2
mg/L after the 5-day incubation period,
and also result in at least 1 mg/L residual
DO (final DO).

Over Mixing the Ployseed

Never let the vortex
touch the stir bar

Micro-organisms in the
seed will be too tired to
get the job done in your
samples and may see
low results in the seed
factor.

Proper way to mix the Polyseed

Mix on a speed of about 5,
or so that the vortex is not
touching the stir bar and
splashing out.
Mix for an hour

Let bran settle out and

transfer to another beaker to
allow to mix for up to 5 hours
on a speed setting between
1-2

Seed Calculations
The DO uptake attributable to the seed (the
seed factor or SF) added to each bottle is
between 0.6mg/L 1.0 mg/L. The SF is
calculated by using the equation below:

SF = (B1 B2) x (f)

Seed Calculations
SF = (B1 B2) x (f)
Where :
B1 = Initial Seed Control DO (before incubation)
B2 = Final Seed Control DO (after 5-day
incubation)
f
= ratio of seed in diluted sample to seed in
seed control , or better see as
f

= (volume, mls of seed in diluted sample)

(volume, mls of seed in seed control)

BOD5, mg/L = (D1 D2) (SF)

P
Where:
D1 = DO of diluted sample
immediately after preparation,
mg/L
D2 = DO of diluted sample after 5-day
incubation period
P = decimal volumetric fraction of
sample used
SF = seed factor

General
Procedure
and
Calculations

General Procedure
300ml BOD bottles are used.
Filled with sample, seed, dilution water to
overfilling (airtight)
Samples brought to room temperature or
20C + 1C before making dilutions

Initial DO is taken before incubation period

(5-days at 20C)
Final DO is taken after incubation period

Unseeded BOD Calculation

BOD5, mg/L = D1 D2
P
Where:
D1 = initial DO of sample
D2 = final DO of sample
P = decimal volumetric fraction of
sample used

EXAMPLE!
150 mLs of a sample was added to a 300
mL BOD bottle and the initial DO of the
sample is 8.2 and the final DO is 4.2, then
what is the BOD5 mg/L?

BOD5, mg/L = D1 D2
P
D1 = 8.2
D2 = 4.2
P = 150 mLs
300 mLs
P = 0.5
BOD5, mg/L = (8.2 4.2)
0.5
BOD5 mg/L = 8 mg/L

SEEDED
BOD CALCULATION
BOD5, mg/L = (D1 D2) (SF)
P
Where D1 = initial DO of sample
D2 = final DO of sample
P = decimal volumetric fraction of
sample used
SF = the DO uptake attributable to
the seed

EXAMPLE!
15 mLs of seed was added to a 300 mL BOD
bottle and labeled as the seed control. The
initial DO was 8.2 mg/L and the final DO is
5.0 mg/L. What is the seed factor, SF if 4
mLs of seed was added to the samples?
Using the calculated SF value, what would be
the BOD5 mg/L if 150 mLs of sample was
added to a 300 mL BOD bottle along with 4
mLs of seed and the initial DO was 8.2 mg/L
and the final DO is 4.2 mg/L?

First Calculate the SF Value

SF = (B1 B2) x (f)

B1 = 8.2
B2 = 5.0
f = 4 mls
15 mls
SF = (8.2 5.0) x 4 mls
15 mls
SF = 0.853

Second calculate the BOD5 mg/L of

the sample:
BOD5, mg/L = (D1 D2) (SF)
P
D1 = 8.2
D2 = 4.2
P = 0.5
SF = 0.853
BOD5, mg/L = (8.2 4.2) 0.853
0.5
BOD5, mg/L = 6.3 mg/L

Interferences

Common Interferences
Samples with caustic alkalinity (pH > 8.5)
Neutralize to a pH between 6.5 7.5

Samples with acidity (pH < 6.0)

Neutralize to a pH between 6.5 7.5

Samples Supersaturated with DO (> 9 mg/L )

Residual Chlorine

Adjusting pH
With samples of high alkalinity or acidity,
use sulfuric or sodium hydroxide solutions
of a concentration of which does not dilute
the sample by more than 0.5%
Standard Methods Recommends a
concentration of 1N acid and alkali solutions
to neutralize caustic or acidic waste samples.

Samples Supersaturated with DO

Samples containing more than 9 mg/L DO
at 20C
Cold water samples
Where photosynthesis has occurred

Reducing Super-saturation
Bring cold samples to room temperature or
20C in a partially filled bottle and agitate
the bottle by shaking vigorously or
aerating with clean filtered compressed
air.

Removing Residual Chlorine

Residual chlorine can cause erroneous
results, where no depletion may occur. In
some samples chlorine will dissipate within
1 to 2 hours of standing in the light. For
samples in which chlorine residual does
not dissipate in a reasonably short time,
destroy the residual chlorine by adding
sodium sulfite (Na2SO3).

Removing Residual Chlorine

De-chlorinating with Sodium Sulfite (Na2SO3)
To the 100 mL sample:

Add 1 mL of a 1:50 sulfuric acid solution

Add 1 mL of potassium iodide solution
2 mL starch indicator
Titrate against 0.025N Na2SO3

X1 = X2

100

150

Where, X1 = known amount of titrant calculated to

neutralize a 100 mL sample
X2 = amount of titrant needed to neutralize 150 mL sample

To calculate the amount of Na2SO3 solution needed to

neutralize a 75mL, 150mL and 250mL sample
X1 = X2
100 75

X1 = X3
100 150

X1 = X4
100 250

(X1)*(75) = (X2)*(100)

(X1)*(150) = (X3)*(100)

(X1)*(250) = (X4)*(100)

(X1)*(75) = X2
100

(X1)*(150) = X3
100

(X1)*(250) = X4
100

Where:
X1 = known amount of titrant calculated to neutralize a 100mL
sample
X2 = amount of titrant needed to neutralize a 75mL sample
X3 = amount of titrant needed to neutralize a 150mL sample
X4 = amount of titrant needed to neutralize a 250mL sample

EXAMPLE #1!
If only using 150 mLs of sample for the BOD
test, and it is known that 3 mLs of 0.025N
Na2SO3 will remove the residual chlorine from
100 mLs of the same sample, then the equation
for how much sodium sulfite the analyst would
need is:

X1 = X2
100 150

X1 = 3 mls
X2 = ?

3.0 = X2
100
150
(3.0)*(150) = (X2)*(100)
(3.0)*(150) = X2
100
X2 = 4.5 mLs 0.025N Na2SO3

EXAMPLE #2!
The residual chlorine in 100 mls of a
sample was removed by titrating with
0.025N Na2SO3 solution. 3 mls of the
titrant was used. Calculate the amount of
titrant necessary to remove the residual
chlorine from 75, 150, and 250 mls of the
same sample.

X1
X2
X3
X4

= 3.0 mls
= amount of titrant needed to neutralize a 75mL sample
= amount of titrant needed to neutralize a 150mL sample
= amount of titrant needed to neutralize a 250mL sample

3.0 = X2
100 75

(3.0)*(75) = (X2)*(100)

3.0 = X3
100 150

(3.0)*(150) = (X3)*(100)

3.0 = X4
100 250

(3.0)*(250) = (X4)*(100)

(3.0)*(75) = X2
100

(3.0)*(150) = X3
100

(3.0)*(250) = X4
100

X2 = 2.25 mls titrant

X3 = 4.5 mls titrant

X4 = 7.5 mls titrant

Determining
Dilutions to Make
on a Unknown
BOD Sample

Determining Dilutions of an
Unknown

A PT Demand Standard has a BOD

concentration range of 15.0 250 mg/L.
What dilutions are needed to find the
result with in this range?

Determining Dilutions of an
Unknown
BOD5, mg/L = D1 D2
(%X)
Where:
DO1 = Initial DO
DO2 = Final DO
X = % volume of sample to be
used

Step #1:

Finding the dilutions on the lower end of the range (15.0 mg/L)

BOD mg/L = 15.0 mg/L

DO1
= 8.0
DO2
= 6.0 (assume a depletion of 2.0)
15.0 mg/L = 8.0 6.0
(%X)
(15.0 mg/L) x (%X) = 2.0
(%X) = 2.0
15.0 mg/L

(%X) = (0.13333) x 100

= 13.33% (round off to 13.3 %) = 40 mLs

Step #2: Finding the dilutions on the lower end of the range (15.0 mg/L)
BOD mg/L = 15 mg/L
DO1
= 8.0
DO2
= 1.0
15 mg/L = 8.0 1.0
(%X)
(15 mg/L) x (%X) = 7.0

(%X) = 7.0
15 mg/L
(%X) = (0.4666) x 100
= 46.666% (round off to 47 %) = 141 mLs or round off to 140 mLs

Finding the dilutions on the lower end of

the range (15.0 mg/L)
Step #1 yielded 13.3% (or 40 mLs sample)
Step #2 yielded 47% (or 141 mLs sample)

Step #1:

Finding the dilutions on the higher end of the range (250.0 mg/L)

BOD mg/L = 250.0 mg/L

DO1
= 8.0
DO2
= 6.0 (assume a depletion of 2.0)
250.0 mg/L = 8.0 6.0
(%X)
(250.0 mg/L) x (%X) = 2.0
(%X) = 2.0
250.0 mg/L

(%X) = (0.008) x 100

= 0.8% = 2.4 mLs (round off to 2.0 mLs)

Step #2:

Finding the dilutions on the higher end of the range (250.0 mg/L)

BOD mg/L = 250 mg/L

DO1
= 8.0
DO2
= 1.0
250 mg/L = 8.0 1.0
(%X)
(250 mg/L) x (%X) = 7.0
(%X) = 7.0
250 mg/L

(%X) = (0.028) x 100

= 2.8% (round off to 3 %) = 9 mLs

Finding the dilutions on the higher end of

the range (250.0 mg/L)
Step #1 yielded 0.8% (or 2.0 mLs sample)
Step #2 yielded 3% (or 9 mLs sample)

Recommended Dilutions from

Standard Methods 20th Edition
0.0 1.0% for strong industrial wastes
1.0 5.0% for raw and settled
wastewaters
5.0 25 % for biologically treated effluents
25 100% for polluted river waters

Questions??

References

American Public Health Association; American Water

Works Association; Water Environment Federation (1998).
Standard Methods for the Examination of Water and Waste
Water. 20th Edition, 4500-O Oxygen (Dissolved)

American Public Health Association; American Water

Works Association; Water Environment Federation (1998).
Standard Methods for the Examination of Water and Waste
Water. 20th Edition, 5210B 5-Day BOD Test
Baird, R.B., & Smith, R.K.P. (2002). Third Century of
Biochemical Oxygen Dmand. Alexandria, VA: Water
Environment Federation.