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BACKGROUND
One of the best medium for growth of microorganism is milk. Both raw milk as well as the pasteurized milk contain varied microorganisms. It is because of the oxygen released by these active microorganisms that the oxidation reduction potential of the milk is decreased. Some of the common microorganism that contaminates milk are staphylococcus aureus, Bacillus Sp. Pseudomonas aeruginosa, Enterobacter sp. and many more. This contaminated milk is one of the biggest source through which diseases are transported from animals to human beings. Mostly milk is contaminated when it is handled properly during milking process when the microorganisms from the exterior surface of the animal comes in contact with the milk. Other source of contaminations is the milk storing utensils like milk cans, and also milking machines. Thus, one of the popular tools for determination of the milk quality is development of Methylene Blue Reductase.
The basic objective of this experiment is to determine the quality of the milk through Methylene Blue reduction test.
Principle: The principle behind this test is that the quality of the milk is determined by observing the colour that appears in the milk after addition of dye like methylene blue. Because methylene blue is a redox indicator, which loses its colour when it comes under the effect of lack of oxygen. Bacterial metabolism causes decrease of oxygen in the milk due to formation of reducing substances. The time taken by the dye to get reduced determines the amount of microbes in the milk and the metabolic reactions in the microorganism.
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REQUIREMENTS
Sample: Milk
Reagent: Methylene blue
Miscellaneous: Screw cap test tubes
Test tube rack
Pipettes
Water bath
Bunsen burner
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PROCEDURE
Take a properly labelled test tube and to it add 10 ml of milk sample. Then to it add 1 ml of methylene blue. Now close the test tube very tightly with screw cap. Now slowly and carefully invert the test tubes for about 4-5 times so that methylene blue mixes with the milk evenly. Now keep the test tubes in water bath at a temperature of 37oC. Now keep a note of the incubation time as it is the time taken for the colour to turn into white colour. Now steady the tube for about 5 minutes.1
A positive result occurs when the viable bacteria present in the milk decolorizes the milk within certain time and when the milk is not decolorized then it is negative result.
If the reduction occurs within 30 min then very poor-quality milk. If reduction occurs between 30 min to 2 h then poor quality milk. When reduction occurs between 2 hours to 6 hours then it is fair quality milk and if reduction occurs between 6 hours to 8 hours then it is good quality milk and finally if not reduced in 8 hours then excellent quality milk.
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CONCLUSION
Methylene blue reduction test gives an accurate result about the quality of the milk and the amount of contaminants present in it.
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REFERENCES
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Thornton HR, Hastings EG. Studies on oxidation-reduction in milk II. The choice of an indicator for the reduction test. The reduction of Janus green b in milk. J Bacteriol. 1929;18(5):319–32.
