COD (Open Reflux)

Methode : Open Reflux
Apparatus
a. Reflux flasks, consisting of 250 mL flask with flat bottom and with 24/29 ground
glass neck
b. Condensers, 24/29 and 30 cm jacket Leibig or equivalent with 24/29 ground glass
joint,or air cooled condensers, 60 cm long, 18 mm diameter, 24/29 ground glass
joint.
c. Hot plate or gas burner having sufficient heating surface.

Reagent
a. Standard potassium dichromate solution, 0.0417M (0.25N): Dissolve 12.259 g
K2Cr2O7,primary standard grade, previously dried at 103oC for 2 hours, in
distilled water and dilute to 1L.
b. Sulphuric acid reagent: Add 5.5g Ag2SO4 technical or reagent grade, per kg of
conc.H2SO4, keep for a day or two to dissolve.
c. Ferroin indicator solution: Dissolve 1.485g 1, 10-phenanthroline monohydrate and
695 mg FeSO4.7H2O in distilled water and dilute to 100 mL. Commercial preparation
may also be available.
d. Standard ferrous ammonium sulphate (FAS), titrant, 0.25M: Dissolve 98g Fe (NH4)2
(SO4)2.6H2O in distilled water, add 20 mL conc. H2SO4, cool and dilute to 1L,
standardise daily as follows.
e. Standardisation: Dilute 10 mL standard K2Cr2O7 to about 100 mL, add 30 mL conc
H2SO4, cool. Add 2 drops of ferroin indicator and titrate with FAS.



g. Mercuric Sulphate, HgSO4, powder
h. Potassium hydrogen phthalate (KHP) standard: Lightly crush and dry potassium
hydrogen phthalate (HOOCC6H4COOK), at 120oC, cool in desiccator, weigh 425 mg in
distilled water and dilute to 1L. This solution has a theoretical COD of 500
μgO2/mL, stable for 3 months in refrigerator.

Procedure
a. Add 50 mL of sample or an aliquot diluted to 50 mL with distilled water in a 500
mL refluxing flask. Add 1g HgSO4, few glass beads, and 5 mL sulphuric acid
reagent, mix,cool. Add 25 mL of 0.0417M K2Cr2O7 solution, mix. Connect the flask
to the condenser Laboratory Manual ID: 1.4 Version: 2 Page: 2/2 and turn on
cooling water, add additional 70 mL of sulphuric acid reagent through open
end of condenser, with swirling and mixing.
b. Reflux for 2 hours; cool, wash down condenser with distilled water to double the
volume of contents, cool.
c. Add 2 drops of Ferroin indicator, titrate with FAS the remaining potassium
dichromate,until a colour change from bluish green to reddish brown. Also reflux
and titrate a distilled water blank with reagents.
d. Use standard 0.00417M K2Cr2O7, and 0.025M FAS, when analysing very low COD
samples.
e. Evaluate the technique and reagents by conducting the test on potassium hydrogen
phthalate solution.
f. Do not add grease at the Leibig jacket to prevent jamming, use water instead.

Calculation

where:
A = FAS used for blank, mL
B = FAS used for sample, mL
M = Molarity of FAS
Note
• Theoretically the method is suitable for analysing samples containing 1000 mg/L COD
without dilution
• In order to economise on quantities of chemicals used in the test, use smaller
sample volumes and proportionally reduce quantities of chemicals as given in the
following table.

Safety Motorcycle Riding

BOD (3 DAYS, 27°C)

Methode : BOTTLE INCUBATION FOR 3-DAYS AT 27°C
Apparatus
a. BOD bottles, 300 mL, narrow mouth, flared lip, with tapered and pointed ground
glass stoppers.
b. Air incubator or water bath, thermostatically controlled at 27 ± 1°C. Light entry
must beprevented in order to avoid photosynthetic oxygen production
c. Accessories: plastic tube, screw-pin and a 5-10 L water container.

Reagents
a. Phosphate buffer solution. Dissolve 8.5 g KH2PO4, 21.75 g K2HPO4, 33.4 g
Na2HPO4.7H2O and 1.7 g NH4Cl in 1L distilled water.
b. Magnesium sulphate solution. Dissolve 22.5 g MgSO4.7H2O in 1L distilled water.
c. Calcium chloride solution. Dissolve 27.5 g CaCl2 in 1L distilled water.
d. Ferric chloride solution. Dissolve 0.25 g FeCl3.6H2O in 1L distilled water.
e. Acid and alkali solution. 1N NaOH and 1N H2SO4. Use for neutralising samples.
f. Glucose-glutamic acid solution (prepare fresh). Dissolve 150 mg dry reagent grade
glucose and 150 mg dry reagent grade glutamic acid in 1L distilled water
g. Sample dilution water. Add 1 mL each of phosphate buffer, MgSO4, CaCl2 and FeCl3
solutions per litre distilled water.

Procedure
a. Prepare required amount of dilution water at the rate of 1000 to 1200 mL per
sample per dilution. Bring the diluted water temperature to 27°C. Saturate with
air by shaking in apartially filled bottle, by bubbling with organic free
filtered air or by storing in cotton-plugged bottles for a day.
b. Some samples do not contain sufficient microbial population (for example, some
industrial wastes, high temperature wastes, or wastes with extreme pH values).
For such wastes, the dilution water is seeded using effluent from a biological
treatment system processing the waste. Where this is not available, use
supernatant from domestic wastewater after settling for at least 1 h but not more
than 36 h. Seed from a surface water body receiving the waste may also be
suitable. Add enough seed volume such that the DO uptake of the seeded dilution
water is between 0.6 and 1.0 mg/L. For domestic wastewater seed, usually 4 to 6
mL seed / L of dilution water is required. Surface water samples usually do not
require seeding.Laboratory Manual ID: 1.2 Version: 1 Page: 2/3
c. Dilution of sample. Dilutions must result in a sample with a residual DO (after 3
days of incubation) of at least 1 mg/L and a DO uptake of at least 2 mgl/L. Make
several dilutions using the Table and experience with the particular sample
source. Polluted surface waters may have 5 to 25 mg/L BOD
Using percent mixture By direct pipetting into 300mL bottles



Table Dilutions for varying BOD values
For preparing dilution in graduated cylinders, siphon dilution water, seeded if necessary,into a 1 to 2 L capacity cylinder. Siphoning should always be done slowly without bubbling,use a screw-pin on the tube to regulate the flow. Keep the tip of the tube just below the water surface as it rises. Fill cylinder half full, add desired quantity of sample and dilute to appropriate level, mix with plunger type mixing rod. Siphon mixed diluted sample in three BOD bottles, stopper without entraining any air. Determine initial DO (method 1.9) on one bottle and incubate the other two at 27°C. Determine final DO (method 1.9) in duplicate after 3days.
For direct pipetting, siphon the desired sample volume to individual bottles and fill with enough dilution water. Complete the test as in the earlier case.

d. Dilution water blank. Find the DO consumption of unseeded dilution water by
determining initial and final DO as in c above. It should not be more than 0.2
mg/L
e. Seed control. Determine the DO uptake by seeding material according to the
procedure in above.

Calculation
a. When dilution water is seeded:


b. When dilution water is not seeded:


where:
D0 = DO of diluted sample initially, mg/L
DT = DO of diluted sample after 3 day incubation at 27°C, mg/L
P = decimal volumetric fraction of sample used
Laboratory Manual ID: 1.2 Version: 1 Page: 3/3
B0 = DO of seed control initially, mg/L
BT = DO of seed control after incubation, mg/L
f = ratio of %seed in diluted sample to %seed in seed control
Notes:
• Report average results of duplicates if both dilutions are correct.
• Formula does not correct for BOD of dilution water which is only valid for
dilution water meeting the criteria. BOD of dilution water should not be more than
0.2 mg/L, preferably lower than 0.1 mg/L.
• The standard glucose-glutamic acid should have BOD of198 ± 37 mg/L (BIS3025 (part
44): 1993). Check procedure otherwise.
• Report BOD values lower than 0.5mg/L or 2 times the measured BOD of the dilution
water (whichever is lower) as lower than detection limit.