Application Information
Apply for laboratory accreditation.
Program Information
More information about the LabCert program.
Resources & Training
Tools to assist you in testing.
Contact information
For information on the Lab Certification program, contact:
Rick Mealy
Program chemist
Science Services / Laboratory Certification

DNR LabCert mailbox

Ultimate BOD Resource

Biochemical oxygen demand

Biochemical oxygen demand (BOD) provides a measure on the impact of a waste (water) on the oxygen content of a receiving system: a stream, river or lake. Wastes are broken down by microbial organisms, frequently referred to as "bugs", who require oxygen for this monumental effort. Thus, in order for this test to work, you need a food source, a nice population of bugs, available oxygen to drive the bugs, and a system which provides a hospitable environment for the bugs.

Bioassay a shorthand form, commonly used term for biological assay and is a type of in vitro experiment. Bioassays are typically conducted to measure the effects of a substance on a living organism. Bioassays may be qualitative or quantitative, the latter often involving an estimation of the concentration or potency of a substance by measurement of the biological response that it produces. Quantitative bioassays are typically analyzed using the methods of biostatistics.

A series of dilutions with nutrient-laden, buffered dilution water is performed on each sample. Samples may also be seeded with a population of microorganisms as necessary. An initial measurement of dissolved oxygen is obtained, and then again following a five-day incubation period at 20 ±1 °C. The extent to which oxygen is depleted is used to calculate BOD. The BOD test is actually a bioassay, it is critical that documentation of conformance with all method parameters is maintained.

Five-day test

There are lots of stories and rationale out there for why BOD is a five-day test, but only one of them is accurate. The BOD test originated in the United Kingdom due to pollution in the London area along the Thames River. Basically, they found that sewage dumped in the Thames took five days to reach the ocean, hence the five-day incubation period. The test was officially adopted in 1908. The Royal Commission on Sewage Disposal, after adopting the BOD test also established the 30 mg/L standard which is used yet today in most permits.

The BOD pyramid

At the heart of BOD testing is a concept that we call the "BOD Pyramid". Just as the food pyramid is the key to maintaining a healthy diet, the BOD Pyramid is the key to producing quality BOD results. Representing the three points of the triangle are the three critical aspects of BOD testing:

  • a food source,
  • a nice population of bugs, and
  • available oxygen to be utilized by the bugs.
the BOD pyramid

The three critical aspects of BOD testing: microorganisms (bugs), oxygen and a food source that requires oxygen for biological degradation. A warm, constant temperature environment in the dark is central.

At the center of the pyramid is the requirement for a hospitable environment for the bugs to thrive.

Virtually all troubles associated with BOD testing can be traced to a disturbance in one of these key aspects, and it often helps to reflect on the importance of the pyramid when troubleshooting your analysis.

Food source

If there is no food source, there can be no BOD, since there would be no oxygen required for breakdown of the food. The logical question to ask is: What if your effluent is ultra-clean; it has no food source for the bugs? The answer is that in very clean samples (200 mLs or more of sample used) we need to add extra nutrients to ensure that bugs continue to survive.

Population of microorganisms (bugs)

On the other side of the coin, we can have a sample with a huge amount of BOD loading, but if there are no living bugs during the BOD test, then no oxygen will be utilized and the BOD results will be unrealistically low. The consequences of this problem would be discharging a waste that may subsequently reduce available oxygen in the receiving stream, having an adverse effect on wildlife downstream.

Available oxygen

You can have bugs and a food source, but there must be oxygen for the bugs to utilize in the biodegradation process. This is one of the reasons why we stress the importance of beginning with a sample that is saturated with oxygen. With a requirement of a minimum depletion of 2 mg/L and a final DO of at least 1 mg/L, we don't have a great deal of working room with which to obtain at least one (two according to 20th ed. Standard Methods) dilution which meets acceptance criteria. If there is not enough oxygen available, then samples over-deplete and we do not have a good measure of the impact of discharging the waste to fish and plant life downstream.

BOD test is not easy

As far back as memory serves, BOD is a test that has — and still continues to be — a test that tends to be assigned to the newest lab analyst or technician. The Wisconsin DNR Laboratory Certification Program establishes fees for test and test categories roughly based on perceived complexity of the testing. The BOD test ranks as the least expensive test category. The lingering question is: If it's such an easy test, why is it associated with the largest number of headaches, phone calls, emails and QC exceedances?

The EPA publication, "Handbook for Analytical Quality Control in Water and Wastewater Laboratories" (EPA-600/4-79-019, March 1979), assigns a skill rating on a scale of 1-4 of 2,3 for BOD.

  • Skill rating 2 - aide with special training or professional with minimum training with background in general laboratory techniques and some knowledge of chemistry, comparable to GS-5 through GS-7.
  • Skill rating 3 - experienced analyst capable of following complex procedures with good background in analytical techniques, professional, comparable to GS-9 through GS-12.

These designations are equivalent to skill ratings associated with performing, among other testing, trace metals analyses.

The reality is that BOD is a tough test. This is not chemistry; this is really a sort of bioassay which is more similar to whole effluent toxicity (WET) testing than any other testing. That being said, the BOD test can be reigned in and kept under control. In order to do that, one has to understand the basic concepts outlined here and on associated links. The basics are: make sure you have happy, healthy bugs and a good calibration to ensure we know precisely how much oxygen was used (accurate initial and final DO readings).

Significance of the BOD test

  • Most commonly required test on WPDES and NPDES discharge permits.
  • Widely used in facility design planning.
  • Assess waste loading on surface waters - what are impacts downstream.
  • Characterized as the "the test everyone loves to hate."

Common problems of the BOD test

  • Sampling and/or preparation issues.
  • Calibration issues.
  • Consistently meeting GGA limits.
  • Ensuring sufficient seed activity.
  • Adding the right amount of seed.
  • DO membrane and probe performance.
  • Sample size.
  • Nitrification.
  • Sample toxicity.
  • Improper interpretation of results.

BOD test limitations

GGA provides flawed metrics

GGA measures accuracy at 200 ppm (which may give an indication of accuracy on influent samples) but most effluents range from 5-25 ppm. How do we know what the accuracy is at these levels which are 1-2 orders of magnitude lower than the GGA level? Additionally, the acceptance criteria for GGA are based on the mean plus/minus a single standard deviation.

  • Test period is too long.
  • Test is not well suited for process control.
  • Test is imprecise and unpredictable.
  • There is no good means to evaluate accuracy - GGA is flawed.
  • No universally accepted standard other than GGA.
  • The test is simply not very easy
    • a lot of QC makes it time-consuming.
    • it can take years of experience to master it.

Alternatives to BOD

Total organic carbon (TOC)
Total organic carbon measures carbon content. All of the carbon in a sample — both organic and inorganic — are oxidized (either chemically or using UV) to carbon dioxide (CO2). Total organic carbon can be provide some measure of the specific pollutants within a waste that are measured as BOD. However, TOC doesn't tell us anything about the amount of organic carbon which is biodegradable (and thus measured as BOD).
Chemical oxygen demand (COD)
Like BOD, COD provides a measure of the amount of organic compounds in water. The difference is that COD is less specific since it measures everything that can be chemically oxidized rather than just levels of bio-degradable organic matter. Chemical oxygen demand also is different in that it reflects the oxidation based on a specific chemical oxidant (dichromate). One final note regarding COD, the EPA approved procedure requires the use of a significant amount of mercury that itself is a pollutant of major environmental concern.

Why BOD is still required

  • None of the alternatives provide a better assessment of the bioavailability of a waste like the BOD test.
  • BOD is a very complex test, however, consistent and reliable BOD results can be produced by any lab if analysts:
    • use good laboratory QC practices;
    • pay attention to details; and
    • carefully follow the approved method.

Copyright 2006. University of Wisconsin Board of Regents.
Unauthorized use prohibited without the expressed written consent of the UW, State Laboratory of Hygiene and the DNR Laboratory Certification & Registration Program.

Last revised: Wednesday August 22 2012