Measuring Dissolved Oxygen using the Winkler Method
DISSOLVED OXYGEN (DO) INTRODUCTION
DO determination measures the amount of dissolved (or free) oxygen present in water
or wastewater. Aerobic bacteria and aquatic life such as fish must have DO to survive.
Dissolved oxygen levels can be negatively affected by pollutants. Microorganisms may
flourish when organic pollutants from wastewater or runoff enter an aquatic
environment. Feeding off of these nutrients, the microbes thrive thereby increasing the
demand of DO which ultimately leads DO decreases.
Dissolved Oxygen is measured in parts per million (ppm).
5-6 ppm Sufficient to most aquatic species
2-4 ppm Stressful to most aquatic species
< 2 ppm Fatal to most species
What is the Winkler Method?
The Winkler Method is a technique used to measure dissolved oxygen in freshwater
systems. Dissolved oxygen is used as an indicator of the health of a water body, where
higher dissolved oxygen concentrations are correlated with high productivity and little
pollution. This test is performed on-site, as delays between sample collections and
testing may result in an alteration in oxygen content.
How does the Winkler Method Work?
The Winkler Method uses titration to determine dissolved oxygen in the water sample.
A sample bottle is filled completely with water (no air is left to skew the results). The
dissolved oxygen in the sample is then "fixed" by adding a series of reagents that form
an acid compound that is then titrated with a neutralizing compound that results in a
color change. The point of color change is called the "endpoint," which coincides with
the dissolved oxygen concentration in the sample. Dissolved oxygen analysis is best
done in the field, as the sample will be less altered by atmospheric equilibration.
Equipment and Material:
Gloves
Safety Goggles
60 mL glass water sampling bottle
Magnesium Sulfate Solution
Alkaline Potassium Iodide Azide
Sulfuric Acid
Sodium Thiosulfate
Starch Indicator Solution
Titrator
Direct Reading Titrator
Thermometer
Procedure:
1. Rinse sampling bottle with the water being sampled
2. Tightly cap the bottle and submerged to desired depth
3. Remove the cap from the bottle allowing bottle to fill. Tap the sides to dislodge
possible air bubbles
4. Replace the cap
5. Remove cap and immediately add:
a. 8 drops of Maganous Sulfate Solution
b. 8 drops of Alkaline Potassium Iodide Azide
6. Cap the bottle and mix by inverting several times. A precipitate will form.
7. Allow the precipitate to settle (below shoulder of the bottle)
8. Add: 8 drops of Sulfuric Acid
9. Cap and gently invert the bottle to mix the contents until the precipitate and
reagent have total dissolved. The solution will be clear yellow to orange if the
sample contains DO.
NOTE: the sample is now Fixed and contact between the air and sample will not affect
the results. This allows you to hold samples for titration at a later point and duplicate
the analysis of a sample.
10. Fill the titration tube the 20 mL line
11. Depress plunger of the Titrator
12. Insert Titrator into the pig in the top of the Sodium Thiosulfate titrating solution
13. Invert the bottle and slowly withdraw the plunger until the large ring on the
plunger is opposite the zero(0) line on the scale
14. Turn the bottle upright and remove the Titrator
15. Insert the tip of the Titrator into the opening of the titration cap
16. Slowly depress the plunger to disperse the titrating solution until the yellowbrown color changes to a very pale yellow. Gently swirl the tube during the
titration to mix the contents.
17. Carefully remove the Titrator and cap. Do not disturb the Titrator plunger.
18. Add: 8 drops of Starch Indicator Solution. The sample should turn blue.
19. Cap the titration tube. Reinsert the tip of the Titrator into the opening of the
titration tube cap.
20. Continue titrating until the blue color disappears and the solution becomes
colorless.
21. Read the test results directly from the scale where the large ring on the Titrator
meets the Titration Barrel. Record as ppm Dissolved Oxygen. Each minor
divisions in the Titrator scale = 0.2 ppm
0.1 mL Sodium Thiosulfate used = 1 ppm Dissolved Oxygen (or 1 mg DO per L of water)
Example: 1.2 mL Sodium Thiosulfate used = 12 ppm Dissolved Oxygen
Water temperature affect the amount of dissolved oxygen an aquatic ecosystem can
hold. To determine the percent saturation, locate the temperature (deg C) of the water
sample on the top scale. Locate the corrected dissolved oxygen concentration on (ppm)
on the bottom scale. Draw a straight line between the two points. Read the % saturation
where the line crosses the % saturation scale.