Biochemical Oxygen Demand (BOD)

The term "seed" refers to microorganisms that consume the biodegradable organic matter in samples for measurement of BOD. Domestic wastewater (influent and effluent) from biological water treatment plants (before disinfection), provide the best source of seed and give the most reproducible results. Other sources, such as industrial wastewater, may not have enough microorganisms or may contain toxins that prevent the organisms from growing. If wastewater is not available, prepare a seed solution from a freeze-dried capsule such as PolySeed ^®. Whatever source of seed is used, it will exert some demand. Therefore, a seed control must be measured to correct for this demand. Older versions of the standard methods for the examination of water and wastewater indicated that dissolved oxygen (DO) uptake of seeded dilution water should be between 0.6-1.0 mg/L; newer versions indicate a minimum of 2.0 mg/L. Refer to the current standard methods or your local regulatory official to verify this requirement.

When determining BOD, it is necessary to have a population of microorganisms that can oxidize (or consume) the biodegradable organic matter present in the sample. If there is too little seed present in the sample, complete consumption of biodegradable matter may not occur, resulting in inaccurate results. In samples such as influent samples and effluent waters prior to disinfection, this is not a problem as the sample will contain sufficient bacteria to do the job. However, in certain sample types (such as some industrial wastes, high temperature wastes and treated effluent), there is not enough bacterial activity to consume the material that is present. In these cases, seed must be added. Seed is simply a solution that contains a sufficient population of bacteria. Hach offers PolySeed ^®, a seed capsule that can be added to samples.

Seed requires proper pH, temperature control and nutrients such as phosphorus, calcium and magnesium for proper growth. Hach nutrient buffer pillows provide the necessary nutrients and pH.

Quality dilution water is very important when testing BOD, as any contamination in the water will cause problems with the test.

Distilled and deionized water are commonly used in BOD testing. However, the most practical way to consistently produce water of low organic content is by distillation with alkaline permanganate. Commercial stills can be set up to produce high-quality distilled water automatically. When a still is fed with chlorinated water, some chlorine may distill over with the water. If this occurs, the chlorine must be destroyed using thiosulfate.

It is not recommended to use deionized water from an ion exchange column. Experience has shown that deionized water, particularly from a new demineralizer with new resin, often contains substantial amounts of organic matter, that is released intermittently and is undetectable with a conductivity water purity gauge. Also, the large surface-to-volume ratio that exists in the columns because of the resin beads, encourages bacterial growth in the column.

For testing BOD, there are several different reasonable sources for seed material. The following are some of the most used:

Plant Influent – If the plant has a stable influent, this is often the most successful seed source. For many plants, the primary influent is reliably a particular value. If the influent is stable and is mostly domestic in origin, consider this as the first seed source.

• It is recommended to settle the seed before use by placing the seed in a covered beaker in an incubator overnight.
• Decant off the particle free seed for use in samples that need seed.

Primary Effluent – Effluent from the primary clarifier is another very good source for seed. It has an advantage over influent in that it is "settled" seed, so most particulates are not present in the seed. This eliminates the settling step.

Final Effluent (prior to disinfection) – If the effluent is sampled prior to disinfection, it may not be necessary to seed the effluent. Many facilities will still need to run glucose and glutamic acid (GGA) standards, so the final effluent is an excellent choice in this case.

• It is recommended to allow the seed to acclimate to room temperature before use. Shake the effluent to help the seed acclimate to room temperature.
• Add 300 mL nutrient pillows to the bottle directly since final effluent may be nutrient deficient.

Artificial Seed – Artificial seed often does not work. If GGA values are low, the artificial seed is most likely the culprit (even if it's been used for years).

• It is recommended to use other types of seed.
• If one must use artificial seed, be sure to follow the vendor's recommendation.
• Seed as high as possible (for example, 1.2 mg/L of depletion).
• Allow as much time as possible for the seed to "wake up" from its freeze-dried state prior to use (but after hydration).
• Add a small quantity of influent or primary effluent to the seed while hydrating it.

Dirt – Often overlooked, soil is a good source of seed material. The bacteria that perform the BOD tests are actually soil bacteria. If none of the other seed sources are adequate, consider experimenting with soil.

• Prepare 500 mL of dilution water (with buffers) in a 1000 mL beaker.
• Place a stir bar into the beaker and keep the water stirred actively.
• Obtain about 20 grams of soil from an actively growing lawn area.
• Place the 20 grams of soil into the 500 mL beaker.
• Allow approximately 1/2 hour of mixing.
• Filter and decant the water into a second 1000 mL beaker.
• Use the solution as a seeding solution.

When testing BOD, standard methods recommend that the seed contribution be 0.6-1.0 mg/L oxygen depletion, though this can be adjusted to ensure GGA standards read 198 +/- 30.5 mg/L.

TOC measures organic carbon but different organic carbons will generate different oxygen demand. Measuring TOC alone will not necessarily indicate how much oxygen will be consumed by the organics in the environment. For example, oxalic acid and ethanol produce identical TOC results. However, due to different oxidation states, the oxygen demand of ethanol is 6 times greater than oxalic acid, meaning ethanol will have a greater effect on the dissolved oxygen content of a receiving water. Measuring oxygen demand rather than TOC produces a clearer picture of how the receiving waters will be affected by the organic-containing wastewaters.