A Citizen's Guide to Understanding and Monitoring Lakes and Streams

Chapter 4 - From the Field to the Lab


How to Measure Dissolved Oxygen

Field Sampling Considerations

Dissolved oxygen concentrations may change drastically in lakes depending upon depth and distance from shore. Sampling stations and depths should be selected according to whether or not you are trying to measure these differences or not. If just one surface station is being measured, pick a station near the middle of the lake and collect the sample at arm’s length below the water surface.

When collecting stream DO samples at several stations for comparison, it is important to select stations with similar flow conditions. Do not select one station in a slow-moving pool and another in a riffle area (unless of course one of your objectives is to measure these differences). The best sites are smooth-flowing – like the "glide" area between riffles and pools.Collecting Samples - Copyright by Sandra Noel

DO samples should represent average conditions in the stream reach being measured. A sample collected in the middle of the stream at least a few inches below the water surface is a safe bet. If the sample must be collected from the shore, be sure to pick a site where there is enough current to ensure adequate mixing – don’t sample from stagnant, slow-moving water if it is not representative of the stream segment.

Assuming your objective is to compare measurements between stations or between seasons, DO samples should be collected at nearly the same time of day each time you sample. Otherwise, the daily variations in DO concentration that were described in Chapters Two and Three may mask changes due to other factors. The time of sampling and water temperature should be recorded. This problem with daily variations in DO (and other parameters) also comes into play if you sample more than one station. For example, if it takes a full day to accomplish the entire monitoring effort, then by default some stations will be sampled in mid-morning, while others will be sampled in mid-afternoon. To retain as much consistency as possible in the data collected, always sample your stations in the same order.

The Use of Field Kits for Water Quality Monitoring

Many of the measurements described in this guide can be made with the use of water quality monitoring "field kits." Forget about test tubes, glass beakers, expensive electronic equipment, or a technician in a white lab coat. These kits are convenient, easy to use, and come with clear, simple directions. The measurements can be quickly made while you are still in the field. This certainly beats the alternative of taking samples into a lab and spending hours doing complex analyses.

However, the difference between a kit and traditional lab techniques is like the difference between opening a can of soup and making your own from scratch. The instant version just does not meet the standards of the traditional method. Likewise, kit measurements do not meet the requirements for precision and accuracy needed for professional quality data. At the same time, kits can play an important role in monitoring programs. Their usefulness is highly dependent upon the monitoring objectives. They are a great educational tool and can provide good broad-based data for general use.

Specific instructions on how to use kits have not been included in this guide because they are provided with the kits and will vary according to the kit manufacturer.

Measurement Methods

There are three common methods for measuring DO. The firsts and most reliable is the Azide-Winkler titration method, against which the others are compared to test for accuracy. However, this method also requires the most training and the use of some strong chemicals. For these reasons, it is not often used in citizen monitoring programs. The second and probably most common method is the use of a DO probe and meter. DO also can be measured with field kits.

For all three methods, the most important step may be the collection of the sample. Precautions must be taken to ensure the sample isn’t aerated during collection and that no bubbles are trapped in the container. Both the Winkler method and kits require that samples be collected into a special type of bottle called a BOD bottle.

If you are hand dipping the BOD bottle, lower the bottle about halfway into the water and let it fill slowly. If you are sampling in a stream, allow the water to overflow for a least 2 minutes or until the water in the bottle has replaced itself two or three times. Check to be sure no air bubbles are present before you lift the bottle – look closely just below the neck of the bottle, where bubbles often get caught. If you see bubbles, gently tip the bottle to either side to allow bubble to escape. Carefully stopper the bottle so no air pockets form below the cap. Do this by tilting the BOD bottle slightly and slowly lowering the cap. You may want to turn the bottle upside down and watch for bubble movement. If you see bubbles, dump the sample and start over.

If the sample was obtained by a sampling device of some kind, the water can not be simply poured into a BOD bottle, since this would cause aeration of the sample. Instead, the sample must be drawn off from a tube located near the bottom of the sampling device. Place the rubber tube into the bottom of the BOD bottle and fill the bottle, again allowing the bottle to overflow until the water has the bottle to overflow until the water has been replaced two or three times. While still letting sample water flow down the tube, slowly pull the tube from the bottom of the bottle and fill the bottle to its brim. Check for bubbles. Carefully stopper the BOD bottle as described above.

Azide-Winkler Method

  1. Fill a 300-mL glass stoppered BOD bottle with sample water. Remember – no bubbles!
  2. Immediately add 2mL of manganese sulfate to the collection bottle by inserting the calibrated pipette just below the surface of the liquid. (If the reagent is added above the sample surface, you will introduce oxygen into the sample.) Squeeze the pipette slowly so no bubbles are introduced via the pipette.
  3. Add 2 mL of alkali-iodide-azide reagent in the same manner.
  4. Stopper the bottle with care to be sure no air is introduced. Mix the sample by inverting several times. Check for air bubbles; discard the sample and start over if any are seen. If oxygen is present, a brownish-orange cloud of precipitate or floc will appear. When this floc has settle to the bottom, mix the sample by turning it upside down several times Dissolved Oxygen Method - Copyright by Sandra Noeland let it settle again.
  5. Add 2 mL of concentrated sulfuric acid via a pipette held just above the surface of the sample. Carefully stopper and invert several times to dissolve the floc. At this point, the sample is "fixed" and can be stored for up to 8 hours if kept in a cool, dark place. As an added precaution, squirt distilled water along the stopper, and cap the bottle with aluminum foil and a rubber band during the storage period.
  6. In a glass flask, titrate 201 mL of the sample with sodium thiosulfate to a pale straw color. Titrate by slowly dropping titrant solution from a calibrated pipette into the flask and continually stirring or swirling the sample water.
  7. Add 2 mL of starch solution so a blue color forms.
  8. Continue slowly titrating until the sample turns clear. As this experiment reaches the endpoint, it will take only one drop of the tritrant to eliminate the blue color. Be especially careful that each drop is fully mixed into the sample before adding the next. It is sometimes helpful to hold the flask up to a white sheet of paper to check for absence of the blue color.
  9. The concentration of dissolved oxygen in the sample is equivalent to the number of milliliters of titrant used. Each milliliter of sodium thiosulfate added in steps 6 and 8 equals 1 mg/L dissolved oxygen.

NOTE: Be very careful when doing DO analyses. The reagents are corrosive, so keep them away from your skin and clothes. Wear safety goggles and wash your hands when you are done.

Probe and Meter Method

  1. Calibrate the probe according to the manufacturer’s suggestions.
  2. Collect the water sample into any appropriate sample container, being careful to avoid aerating the sample as describe above.
  3. Place the probe in the sample, allow the meter to equilibrate, and read the DO concentration directly off the scale. NOTE: The probe may need to be gently stirred to aid water movement across the membrane.

Field DO probes are easily ruined through deterioration of the membrane, trapping of air bubbles under the membrane, and contamination of the sensing element. It often is difficult to assess whether or not a probe is functioning properly. Because of this, the meter must be calibrated before and after each series of measurements. When you calibrate the instrument, you compare DO concentrations measured by the probe to those measured using the Azide-Winkler method described above and then correct all samples for any measurement error. The meter manufacturer’s calibration procedure should be followed exactly. If the error is high or erratic, all sample results should be discarded.

QA/QC Considerations

Even though the Winkler dissolved oxygen method is the method against which the others are calibrated, there are still tests that can be made to ensure that the Winklers themselves are accurate. To test the method, you need to have samples with a known oxygen concentration so you can compare your results to what you know is the real answer. These are called calibration samples or standards. A 100 percent saturation solution can be prepared by bubbling air into distilled water. If low DOs are expected, a zero DO solution can be made by adding excess sodium sulfite and a trace of cobalt chloride to a sample. In a professional lab, a calibration standard would be analyzed with each bath of samples run.

Randomly select 5 to 10 percent of the samples for duplicate laboratory analysis. If you are interested in field variability, select 5 to 10 percent of the samples for field duplication (e.g., collect two samples from the same station).

If you are using a probe and meter or field kit for measurement, 5 to 10 percent of your samples should be checked against the Winkler DO method.

The next section discusses how to measure pH.


Measuring Temperature | Measuring Dissolved Oxygen | Measuring pH | Measuring Secchi Disk Depth | Measuring Nutrient Concentrations | Measuring TSS | Measuring Chlorophyll a | Measuring Fecal Bacteria

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