Methods for Analyzing Organic Matter in Water Bodies - Biochemical Oxygen Demand (Bod) - Five-Day Incubation Method

Biochemical oxygen demand (BOD) refers to the amount of dissolved oxygen consumed by aerobic microorganisms during the biochemical oxidation process of decomposing organic matter in water under conditions of dissolved oxygen. It also includes the oxygen consumed by the oxidation of reducing inorganic substances such as sulfides and ferrous oxide, but this usually constitutes a small proportion.

The aerobic decomposition of organic matter by microorganisms generally occurs in two stages. The first stage is called the oxidation stage, mainly where carbon-containing organic matter is oxidized into carbon dioxide and water. The second stage is called the nitrification stage, mainly where nitrogen-containing organic compounds are decomposed into nitrite and nitrate by nitrifying bacteria. However, these two stages are not completely separate, but rather each has its own primary and secondary role. For domestic sewage and industrial wastewater with similar properties, the nitrification stage only becomes significant after approximately 5-7 days, or even more than 10 days. Therefore, the 20°C five-day culture method (BODs method) widely used domestically and internationally for determining BOD values generally does not include the nitrification stage. BOD is a comprehensive indicator reflecting the degree of organic pollution in water bodies. It is also an important parameter in studying the biodegradability and biochemical treatment effect of wastewater, as well as in the design and kinetic research of biochemical wastewater treatment processes.

Five-Day Incubation Method (20℃)

Also known as the standard dilution method. The principle is that after dilution, the water sample is incubated at 29±1℃ for 5 days, and the dissolved oxygen content before and after incubation is calculated. The difference between the two is BOD5. If the five-day biochemical oxygen demand of the water sample does not exceed 7 mg/L, dilution is unnecessary, and it can be measured directly. Many relatively clean river waters belong to this category.

For industrial wastewater containing few or no microorganisms, such as acidic wastewater, alkaline wastewater, high-temperature wastewater, or chlorinated wastewater, inoculation should be performed when measuring BODs to introduce microorganisms capable of degrading organic matter in the wastewater. When the wastewater contains organic matter that is difficult to be degraded at a normal rate by microorganisms in general domestic sewage, or contains highly toxic substances, acclimatized microorganisms should be introduced into the water sample for inoculation.

1. Dilution Water

For contaminated surface water and most industrial wastewater, which contain a high amount of organic matter, dilution is necessary before incubation and measurement to ensure sufficient dissolved oxygen during the incubation process. The dilution level should ensure that the dissolved oxygen consumed during incubation is greater than 2 mg/L, while the remaining dissolved oxygen is above 1 mg/L.

Dilution water is generally prepared using distilled water. First, air treated by activated carbon adsorption and water washing is introduced and aerated for 2-8 hours to bring the dissolved oxygen in the water close to saturation. Then, it is left to stand at 20°C for several hours. Before use, a small amount of nutrient salt solutions such as calcium chloride, ferric chloride, and magnesium sulfate, as well as a phosphate buffer solution, are added and mixed thoroughly. The pH of the dilution water should be 7.2, and the BOD5 should be less than 0.2 mg/L.

If the water sample contains no microorganisms, microorganisms should be inoculated into the dilution water. This involves adding 1-10 mL of supernatant from domestic sewage, 20-30 mL of surface soil leachate, or 10-100 mL of river or lake water per liter of dilution water. This water is called inoculation dilution water. To check the quality of the diluted aqueous inoculum and the operational skill of the laboratory personnel, a standard solution containing 150 ml/L glucose and glutamic acid was diluted at a ratio of 1:50 and simultaneously measured with the water sample for BODs. The measured value should be between 180 and 230 ml/L; otherwise, the cause should be investigated and corrected.

2. Water Sample Dilution Ratio

The water sample dilution ratio should be estimated based on practical experience. Table 2-13 lists the methods for estimating the dilution ratio of surface water. The dilution ratio of industrial wastewater is obtained by multiplying the CODcr value by coefficients of 0.075, 0.15, and 0.25, respectively. Usually, water samples at three dilution ratios are prepared simultaneously. Table 2-13: Coefficients for estimating the dilution ratio using the permanganate index.

3. Calculation of Measurement Results

For water samples directly cultured without dilution:

Where Icl—the concentration of dissolved oxygen in the water sample before culture (ml/L);

: —the remaining dissolved oxygen concentration in the water sample after 5 days of culture (ml/L). For diluted water samples after incubation:

Where: B1—Dissolved oxygen concentration in the dilution water (or inoculum dilution water) before incubation (m³/h);

Bz—Dissolved oxygen concentration in the dilution water (or inoculum dilution water) after incubation (m³/h);

f1—Proportion of dilution water (or inoculum dilution water) in the culture medium;

f2—Proportion of water sample in the culture medium.

When water samples contain toxic substances such as copper, lead, zinc, cadmium, chromium, arsenic, and cyanide, they inhibit microbial activity. Dilution water inoculated with acclimatized microorganisms can be used, or the dilution factor can be increased to reduce the impact of the toxic substances. If a small amount of chlorine is present, it will generally disappear on its own after 1-2 hours. For water samples where free chlorine cannot dissipate in a short time, sodium sulfite can be added to remove it; the amount added should be determined experimentally.

This method is suitable for determining water samples with a BOD5 greater than or equal to 2 m³/h, and a maximum of 6000 m³/h. For BOD5 greater than 6000 m³/h, dilution will introduce greater errors.