Millon’s Test

Millon’s Test for Tyrosine Definition

Millon’s test is specific to detect presence of amino acid Tyrosine.

Millon’s test is a biochemical test used to detect the presence of tyrosine in a given sample. It is named after the French chemist Auguste Millon.

The test involves the addition of Millon’s reagent, which is a solution of mercuric nitrate, nitric acid, and nitrous acid, to the sample being tested. The reagent reacts with tyrosine, producing a reddish-brown color.

Principle of Millon’s Test

The reaction is dependent on the formation of colored Hg- compound with phenolic (OH) group. The reaction is not specific for proteins however tyrosine is the only phenolic amino acid in proteins; hence a positive test indicates the presence of tyrosine. It works by adding a solution called Millon’s reagent to the sample, which reacts with the tyrosine to produce a reddish-brown color. This happens because tyrosine has a special group that reacts with the chemicals in the Millon’s reagent to form a colored complex. This test is often used to identify the presence of tyrosine in proteins.

Requirements for Millon’s test

Chemicals and Reagents

  • 1 ml of the test solution
  • 1 ml of Millon’s reagent
  • Few drops of 5% NaNO2 solution

Instruments and Materials

  • Test tubes
  • Test tube stand
  • Pipettes or Dropper
  • Hot Water bath


  1. Take a dry and sterile test tube.
  2. Add 1 ml of the given sample solution into the test tube with the help of dropper.
  3. Add 1 drop of millon’s reagent in it and shake well.
  4. A yellow precipitate sticking to the sides of the test tube develops due to the precipitation of proteins.
  5. Add a drop of 5% NaNO2 solution.
  6. Observe the change in colouration or precipitation.
  7. Development of a red precipitate or coloration indicates a positive test.

Results and Interpretations of Millon’s Test

The result of Millon’s test is the appearance of a reddish-brown color when Millon’s reagent is added to the sample being tested. The intensity of the color can vary depending on the amount of tyrosine present in the sample.

The interpretation of the test result depends on the context in which it is being used. If the test is being used to identify the presence of tyrosine in a protein, a positive result indicates that the protein contains one or more tyrosine residues. However, it is important to note that other amino acids, such as tryptophan and phenylalanine, can also give a positive result with Millon’s test because they also contain a phenolic group.

Potential Uses of Millon’s Test

Millon’s test is a commonly used biochemical test in various fields, including biochemistry, food science, and medical diagnostics. Some of its main uses are:

  1. Protein identification: Millon’s test is often used to identify the presence of tyrosine residues in a protein. This can be useful in protein sequencing and characterization.
  2. Food analysis: The test can be used to identify the presence of tyrosine in food products, such as milk, cheese, and meat. This information can be used to assess the quality of the food or to detect adulteration.
  3. Medical diagnostics: Millon’s test can be used in the diagnosis of certain medical conditions. For example, a positive result for tyrosine in urine can indicate the presence of a genetic disorder called tyrosinemia.
  4. Chemical analysis: The test can also be used as a general test for the presence of phenols in a sample.

Limitations and Drawback of Millon’s Test

Although Millon’s test is a useful biochemical test, there are some limitations to its use:

  1. Lack of specificity: Millon’s test is not specific to tyrosine and can also give a positive result for other compounds that contain a phenolic group, such as tryptophan and phenylalanine. Therefore, the test must be used in conjunction with other tests to confirm the presence of tyrosine.
  2. Lack of sensitivity: Millon’s test may not be sensitive enough to detect low concentrations of tyrosine in a sample. Therefore, other tests with higher sensitivity may need to be used in conjunction with Millon’s test to accurately detect low levels of tyrosine.
  3. Interference from other substances: Some substances in the sample being tested, such as ascorbic acid or other antioxidants, can interfere with the reaction between Millon’s reagent and tyrosine, leading to false negative results.
  4. Toxicity of reagent: Millon’s reagent contains mercury, which is toxic. Therefore, appropriate safety precautions must be taken when handling the reagent.

FAQs on Millon’s Test

Which proteins give positive Millon’s test?

Millon’s test is given by any compound containing a phenolic hydroxy group which is used to detection of protein Tyrosine.

What is positive in Millon’s test?

Any protein containing tyrosine will give a positive test of a pink to dark-red colour.

References and Sources

Further Readings

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  2. Aseptic Transfer Technique
  3. Bacterial Flagella, Fimbriae and Pili
  4. Growth Curve of Bacteria
  5. Instruments Used in Microbiology Laboratory
  6. MacConkey agar
  7. Monochrome Staining
  8. Negative Staining
  9. Serial Dilution in Microbiology
  10. Spread Plate Technique
  11. Streak Plate Technique