ORGANIC CARBON
(WALKLEY-BLACK TITRATION AND
COLORIMETRIC METHOD)
Ma. Joerdette N. Jimenez
Chemist I, Laboratory Services Division
Bureau of Soils and Water Management
Introduction
Soil carbon is
probably the most
important component
in soils as it affects
almost all soil
properties.
Introduction
PHYSICAL
CHEMICAL
BIOLOGICAL
Introduction
• Better crop yields
• Reduces soil erosion
• Increases biological
diversity
• Increases plant
nutrient retention
Introduction
Increases in soil
organic carbon
enhances the
biomass and
diversity of the soil
biodata.
Introduction
The content of organic
carbon of mineral horizons
can be estimated from the
Munsell color of dry and/or
moist soil, taking the
textural class into account.
Scope and Field of Application
This protocol applies to the determination
of the Oxidizable Organic Carbon content
in soil. This test method does not
routinely apply correction for chloride.
Principle
POINTS TO BE NOTED:
• Walkley and Black found that on the average about 77% of the organic C
was recovered by the heat of dilution procedure, a correction factor of 1.3 be
used to account for unrecovered organic C;
• Chloride, ferrous iron and higher oxides of Mn have been shown to undergo
oxidation-reduction reactions in chromic acid mixtures leading to incorrect
values for organic carbon;
• Smaller sample weights should be used for samples with very high carbon
content;
• It is not applicable to soils containing significant amounts of carbonized
materials.
Principle in Titration Method
6 Fe2+ + Cr2O72- + 14 H+ 2 Cr3+ + 6 Fe3+ + 7 H20
+ =
The organic C can then be estimated by measuring the remaining
unreduced dichromate by back-titrating with ferrous sulphate or
ammonium ferrous sulphate using diphenylamine or o-
phenanthroline-ferrous complex as an indicator.
Apparatus (Titration Method)
❑Analytical Balance
❑Burette, 50 mL
❑Volumetric burette/dispenser, 10 mL
❑Volumetric dispenser
❑Erlenmeyer flasks, 500 mL
❑Magnetic stirrer and bar
❑Oven (105° C)
❑Volumetric flasks, 1000 mL
❑Glass rod
❑Beaker, 100 mL, 250 mL
❑Fume hood
❑Burette stand
Materials (Titration Method)
1. Deionized water/distilled water, it should have an EC < 1.5 *10^-3 dS m^-1
2. Potassium dichromate, 0.167 M (10 N)
3. Sulfuric Acid, Concentrated (not less than 96%)
4. Phosphoric Acid, 85% (If Diphenylamine indicator is used)
5. Indicator (either 5.1 or 5.2)
5.1. o-Phenanthroline – Ferrous Complex, 0.025 M
5.2. Barium diphenylamine sulfonate Indicator, 0.16% aqueous solution
6. Titrant (either 6.1 or 6.2)
6.1. Ferrous Sulphate (FeSO4) solution, 0.5 M
6.2 Ferrous Ammonium Sulphate, 0.5 M
Procedure (Titration Method)
1. Weigh 1.0 g of air-dried soil 6. Add 200 mL of water to the flask
2. Add 10 mL of 0.167 M K2Cr2O7 2- 7. Add 10 mL of 85% H3PO4 (if barium
diphenylamine sulfonate indicator is used)
3. Swirl the flask gently to dispense the soil in 3. Add 3-4 drops of o-phenanthroline or barium
the solution. diphenylamine indicator.
4. Add 20 mL of concentrated sulfuric acid, 4. Titrate the solution with 0.5 M FeSO4
directing the stream into the suspension solution or 0.5 M (NH4)2Fe(SO4)2 * 6H2O
5. Immediately swirl the flask gently until soil 5. Observe for color change from blue to red
and reagents are mixed. (maroon color in white background)
Calculation (Titration Method)
% Organic Carbon = (V blank – V sample) x M Fe x 0.003 x 100 x f x mcf
W
Where:
V blank = volume of titrant in blank, mL
V sample = volume of titrant in sample, mL
0.003 = carbon oxidized
f = correction factor, 1.3
W = weight of soil, g
mcf = Moisture correction factor
Principle in Colorimetric Method
2 Cr2O7 2- + 3 C0 + 16 H+ 4 Cr3+ + 3 CO2 + 8 H20
+ =
Organic carbon can be calculated from the amount of
chromic ion formed, using a colorimetric procedure.
Principle (Colorimetric Method)
• Walkley & Black chromic acid
wet oxidation method.
• OC is oxidized by 0.167 M
potassium dichromate solution
in concentrated sulfuric acid.
Figure 1. Overview of how particulate and mineral associated organic matter form and function.
Retrieved from https://theconversation.com/soil-carbon-is-a-valuable-resource-but-all-soil-carbon-is-not-created-equal-129175
Principle (Colorimetric Method)
• Green chromic ions (Cr 3+)
produced from the reaction is in direct
proportion to the carbon oxidized
which is measured by colorimetric
estimation.
• Oxidation can be determined by
measuring the concentration at
wavelength near 600 nm.
Apparatus (Colorimetric Method)
❑Analytical Balance
❑Spectrophotometer
❑Centrifuge tubes/glass conical tubes,
50 -70 mL
❑Dispensing or volumetric pipettes
❑Graduated pipettes
❑Calibrated dispenser
❑Glass rod
❑Volumetric flasks
❑Beaker
Equipment and Laboratory Ware
UV-Vis Dispenser
Spectrophotometer
Conical
Analytical Tube
Balance
Materials (Colorimetric Method)
1. Deionized water/distilled water, it should have an EC < 1.5 *10^-3 dS m^-1
2. Potassium dichromate, 10% (0.34 M)
3. Sucrose Standard, 4mg/mL
4. Sulfuric Acid, Concentrated (not less than 96%)
Procedure (Colorimetric Method)
PREPARATION OF STANDARDS PREPARATION OF SAMPLES
1. Place volumes of sucrose 1. Weigh soil sample according to
standard and DI water in Table 1. Table 2.
2. Add 2 mL of 10 % Potassium 2. Add 2 mL of 10% Potassium
Dichromate and mix. Dichromate and mix.
3. Add 5.0 mL of Sulfuric acid, cool 3. Add 5.0 mL of Sulfuric acid, cool
and stand for 30 mins. and stand for 30 mins.
4. Add 18 mL DI water. Mix. Stand 4. Add 20 mL DI water. Mix. Stand
overnight. overnight.
5. Read absorbance set at 600 nm. 5. Read absorbance set at 600 nm.
Procedure (Colorimetric Method)
TABLE 1. STANDARD PREPARATION
Mass of OC (mg) Sucrose Standard (mL) Deionized Water (mL)
0 0.00 2.00
1 0.25 1.75
2 0.50 1.50
3 0.75 1.25
4 1.00 1.00
5 1.25 0.75
6 1.50 0.50
7 1.75 0.25
8 2.00 0
Procedure
TABLE 2. Recommended weight of sample.
Weight (g) OC, % Color
0.1 >2 Black, dark gray,dark brown
0.25 ≤0.2 Brown-dark brown, gray-dark gary
0.5 <0.6 brown
Note: Above is just a guide in determining the appropriate weight to be used for each sample based on soil color. % OC may vary
per soil color type. Generally, dark colored soils which are described as dark brown to black show a higher content of carbon and
nitrogen than those soils which are lighter in color.
Calculation (Colorimetric Method)
% Organic Carbon = mg C (sample) – mg C blank x mcf x f x 100
W, mg
% Organic Matter = % O.C. x 1.72
Where:
% O.C. = Organic carbon content of the soil, %
mg C sample = Analyte/concentration of C in sample
mg C blank = Analyte/concentration of C in blank
W = Mass of air-dry sample, mg
mcf = moisture correction factor
f = Correction factor, 1.3
Health and Safety
✔ Safety glasses, gloves and lab
coats must be worn when handling
any chemicals
✔ Potassium Dichromate: Highly
corrosive and is a strong oxidizing
agent.
✔ Sulfuric Acid: Keep away from
naked flames/heat. Always add the
acid to the water.
Quality Assurance/Quality Control
ACCURACY TEST
• Participate in Interlaboratory Proficiency Testing
Program at least once a year.
• Analyze CRM or QRM.
Quality Assurance/Quality Control
PRECISION TEST
Perform replicate analysis at most in every 10% samples of
a batch. Calculate the % RSD and compare the result with the target
precision for the analyte concentration.
Quality Assurance Procedure
CONTROL CHART
Analyzed at least duplicate
of the Check Sample
or Internal Reference Material for every batch of
analysis. Plot result in control chart.
References
AOAC. 1998. AOAC Peer Verified Methods Program. Manual on Policies and Procedures. AOAC
International Gathersburg. MD.
B. Magnusson & U. Ornemark. 2014. Eurachem Guide: The fitness for Purpose of Analytical
Methods – A Laboratory Guide to Method Validation and Related Topics.
Bowman, R.A. 1998. A Re-evaluation of the Chromic Acid Colorimetric Procedure for Soil Organic
Carbon. Commun. Soil Sci. Plant Anal., 29(3&4): 501-508.
Brown, P.E. & O'Neal, A.M. 1923. The Color of Soils in Relation to Organic Matter Content.
Research Bulletin No. 75. Retrieved from Agricultural Research Bulletin-v005-b075.pdf.
FAO. 2006. Guidelines for soils description. Fourth edition. Food and Agriculture Organization of
the United Nations, Rome, Italy.
References
Garfield, F.M. 1991. Quality Assurance Principles for Analytical Laboratories. AOAC
INTERNATIONAL
Nelson, D.W. & Sommers, L.E. 1996. Total Carbon, Organic Carbon and Organic Matter. In D.L.
Sparks (Ed.), Soil Science Society of America, Book Series 5. Methods of Soil Analysis Part 3,
Chemical Methods. Madison, Wisconsin: Soil Science Society of America, Inc.
Rayment, G.E. & Lyons, D.J. 2011. Soil Chemical Methods - Australasia. CSIRO publishing,
Australia
Sims, J. & Haby, V. 1971. Simplified Colorimetric Determination of Soil Organic Carbon Matter.
Soil Science, 112(2): 137-141
References
Walkley, A. & Black I.A., 1934. An examination of the Degtjareff Method for Determining Soil
Organic Matter, and a proposed Modification of the Chromic Acid Titration Method. Soil Science,
37(1): 29-38
Walkley, A. 1947. A Critical Examination of a Rapid Method for Determining Organic Carbon in
Soils – Effect of Variations in Digestion Conditions and of Inorganic Soil Constituents. Soil
Science, 63(4): 251-264
Thank you for
listening!!!
