Determination of Biological Oxygen Demand (BOD) in Waste Water : Pharmaguideline
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  • Jun 9, 2013

    Determination of Biological Oxygen Demand (BOD) in Waste Water

    Determination of the Biochemical Oxygen Demand (BOD) in polluted waste water as a pollution indicator by incubating the BOD bottles for five days at 20 degree C..

    Biological Oxygen Demand (BOD)

    The amount of oxygen, taken up by the microorganisms that decompose the organic waste matter in wastewater is known as biological oxygen demand or biochemical oxygen demand.
    Therefore it is used to measure the amount of certain types of organic water pollution  BOD is calculated by keeping a sample of water containing a known amount of oxygen for five days at 20 °C. 

    The oxygen content is measured again and BOD is calculated. A high BOD indicates the presence of a large number of microorganisms which indicates a high level of pollution in wastewater.

    BOD determination

    1. Neutralization of Sample:

    The accuracy of BOD test totally depends upon the proper bacterial growth, present in the water sample. pH of the diluted sample should be adjusted 7.00 ±0.2 before the incubation for five days for proper results. The sample should be neutralized in the following manner.
    1.1  Take 50 ml of sample in a 100 ml beaker.
    1.2  Measure the pH of the solution by using the calibrated pH meter.
    1.3  Add the 1N sulfuric acid to adjust the pH if it is higher than 7.00 and 1N sodium hydroxide if pH is lower than 7.00.
    1.4  Note down the volume of sulfuric acid or sodium hydroxide used to adjust the pH of 50 ml sample to 7.00 ±0.2.
    1.5  Calculate the volume of sulfuric acid or sodium hydroxide required to neutralize the 1000 ml sample.
    1.6  Add the calculated volume of sulfuric acid or sodium hydroxide to the sample to neutralize.
    For example, if 2.1 ml of 1N sulfuric acid or sodium hydroxide are used to neutralize 50 ml of sample to pH 7.00 ±0.2.  Calculate the volume of 1N sulfuric acid or sodium hydroxide to be added to neutralize the 1000 ml sample as follows:
    1N sulfuric acid or sodium hydroxide required = (2.1 ml x 1000 ml)/50 ml = 2100/50 = 42 ml.
    Note: Hydrochloric acid or other acid containing chlorine should not be used to neutralize the sample because chlorine interferes the results of wastewater BOD.

    2. Removal of Chlorine Content:

    Chlorine is a strong oxidizing agent and it can inhibit the microbial growth during wastewater BOD analysis, so it should be removed from sample before start the analysis. Chlorine can be removed by adding the sodium sulfite to the sample in following manner.
    2.1  Take 50 ml of water sample to be tested in a conical flask.
    2.2  Add 2.5 ml of acetic acid diluted to 50% with water.
    2.3  Add 2.5 ml of 10% w/v solution of potassium iodide.
    2.4  Add 1 ml of starch indicator and titrate with 0.025N sodium sulfite solution.
    2.5  Note down the volume and calculate to add in 1000 ml of the sample as described above in Neutralization of Sample section.
    2.6  Add the calculated volume of sodium sulfite solution to the sample and mix well to neutralize the chlorine.

    3. Preparation of Phosphate Buffer Solution:

    Dissolve accurate weighed 8.5 gm of potassium dihydrogen phosphate (KH2P04), 21.75 gm of Dipotassium hydrogen phosphate (K2HP04), 33.4 gm of Disodium hydrogen phosphate (Na2HP04.7H20) and 1.7 gm of ammonium chloride (NH4Cl) in 500 ml distilled water. Dilute the solution up to 1000 ml.

    4. Preparation of Alkali-Iodide-Azide Reagent:

    Dissolve 500 gm of sodium hydroxide (NaOH) and 135 gm of sodium iodide (NaI) in distilled water. Make up the solution to 1000 ml of distilled water. Now dissolve 10 gm of sodium azide in this solution.

    5. Preparation of Dilution Water:

    The dilution water for wastewater BOD analysis must be free from organic content. Dilution water can be prepared by the following method.
    5.1  Take five liters of double distilled water in a glass container  
    5.2  Aerate the water with clean compressed air for not less than 12 hours.
    5.3  Allow to stable for at least 6 hours at 20 °C.
    5.4  Add 5 ml of 27.5% w/v solution of calcium carbonate.
    5.5  Add 5 ml of 22.5 % w/v solution of magnesium sulfate.
    5.6  Add 5 ml of 0.15% w/v solution of ferric chloride.
    5.7  Add 5 ml of phosphate buffer solution.
    5.8  Mix well and allow to stand for 2 hours.

    6. Procedure to Determine the Biological Oxygen Demand of Water:

    6.1  Take four 300 ml BOD bottles and add 10 ml of samples to two bottles and fill the remaining volume with dilution water.
    6.2  Fill the remaining two BOD bottles only with dilution water for blank.
    6.3  Immediately close the bottles when filled and there should not be any air bubbles in the bottle.
    6.4  Mark the bottles as blank and sample.
    6.5  Incubate one sample and one blank bottle at 20 °C for 5 days.
    6.6  Analyze immediately the remaining one blank and one sample bottle of dissolved oxygen (DO).
    6.7  Analyze incubated bottles for DO after 5 days.

    Recommended sample volume for BOD determination
    BOD range (mg/liter)
    Sample volume (ml)
    Dilution water (ml)

    7. Test for Dissolved Oxygen (DO):

    7.1  Add 2 ml of 36.4% of manganous sulfate (MnSO4.H2O) solution inserting the tip of pipette tip into the sample because the drops of solution can allow inserting the oxygen into the solution.
    7.2  Add 2 ml of the alkali-iodide-azide reagent by above method.
    7.3  Allow reacting the solutions with the oxygen present in the sample.
    7.4  When precipitates are settled down at the bottom add 2 ml of concentrated sulfuric acid by placing the pipette tip very near to sample surface.
    7.5  Mix well to dissolve the precipitates.
    7.6  Take 203 ml of sample from BOD bottle into an Erlenmeyer flask.
    7.7  Titrate immediately with 0.025N sodium thiosulfate solution using starch indicator until blue color disappears and note down the burette reading.
    7.8  Determine the burette reading for blank in the same manner.

    BOD formula and Calculation:

    Blank correction = B.R. for blank at D0 – B.R. for blank at D5
    BOD mg/l = [(B.R. for sample at D0 –D5)– blank correction] x dilution factor
    Dilution factor = Bottle volume (300 ml)
                                   Sample volume
    B.R. = burette reading
    D0 = Initial
    D5 = Day five after incubation

    Also see: Purified Water System Validation

    Frequently Asked Questions on Biological Oxygen Demand

    Q: What is Biological Oxygen Demand (BOD)?

    Biological oxygen demand is the amount of oxygen required by the microorganisms in wastewater to decompose the organic matter. The level of pollution in any water body is estimated by BOD. A higher BOD value indicates greater pollution in the water body.

    Q: Why is BOD important in assessing water quality?

    Determination of BOD is important in assessing water quality because it provides information regarding the pollution in water by organic matter. High BOD values show the higher organic matter dissolved in water that decreases the dissolved oxygen value in the water body and harms the aquatic life.

    Q: How is BOD measured?

    BOD is measured by conducting the biological oxygen demand test, in which the water sample is incubated in special BOD bottles for 5 days at 20° C. During this period microorganisms decompose the organic matter present in water and consume the dissolved oxygen. BOD is calculated by determining the dissolved oxygen in the sample before and after incubation. As described above in detail.

    Q: What are the units of measurement for BOD?

    BOD is measured in mg/liter that is equivalent to parts per million (ppm)

    Q: What factors influence BOD levels in water?

    Factors like organic content, temperature, pH, nutrition availability and the presence of microorganisms can influence the BOD level in water bodies.

    Q: What are the sources of organic pollution that contribute to BOD?

    Domestic sewage is the major source of BOD in water bodies. Other sources like agriculture runoff, industrial wastewater, animal waste and organic matter from decaying plants also contribute to BOD.

    Q: How is BOD used in wastewater treatment?

    BOD level is a primary indicator of water pollution during wastewater treatment. It helps to determine the effectiveness of the treatment process. ETP operators measure the BOD of inlet water and treated water to determine the effectiveness of the wastewater treatment process.

    Q: What is the difference between BOD and COD (Chemical Oxygen Demand)?

    BOD and COD both are used to measure the organic pollution in the water. BOD measures the organic pollutants that are biodegradable while COD measures the whole organic matter either biodegradable or non-biodegradable.

    Q: Are there regulatory standards for BOD levels in water?

    Yes, government authorities and environmental agencies set the acceptable BOD levels for different types of water as drinking water, aquatic ecosystems and treated water.

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    1. Thank you, Good and very much helpful content


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