Laboratory Test of Proteins

Dhristi JEE 2022-24

An introduction to Proteins and Laboratory Test of Proteins

Proteins are important biological compounds that have a higher molecular weight. They are probably nature’s most complex organic materials. Plants tend to build their proteins from minerals, carbon dioxide, and water in the presence of sunlight. These compounds are the most abundant organic compounds found in nature. In every living organism, they are present. Animals tend to get their share of proteins from plants. Life is linked with the existence of proteins. 


The proteins are identified by some laboratory tests. These tests are discussed below along with its theory, required material for tests, and observation. The aim of the protein laboratory test is discussed below.


Aim

The aim of this article to perform a total protein lab test for identifying the presence of proteins in the samples given.


Theory

Proteins possess a higher molecular mass of long-chain polymers that consist of α-amino acids. Cells contain proteins and they are therefore present in all living bodies. Proteins consist of carbon, oxygen, nitrogen, hydrogen, and often sulphur and phosphorus. There are a few tests that are carried out for finding the presence of proteins in the samples given. 


These laboratory tests of proteins are:

  1. Biuret test

  2. Millions test

  3. Xanthoproteic test

  4. Ninhydrin test


Each of the tests is discussed below along with the chemical reaction involved in the test.


  1. Biuret Test

The compounds having peptide linkage undergo the Biuret test. Proteins are known as the polypeptides of amino acids that are linked together by the peptide bonds. A protein alkaline solution is treated with one drop of aqueous copper sulphate and a bluish violet colour is seen.


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The Biuret test is useful in the identification of proteins and protein estimation. Biuret is a kind of chemical that is formed when urea gets heated to 180 ℃. During this reaction, two molecules of urea tend to condense and form a biuret or a bi-urea molecule. The biuret reagent forms a complex of violet colour in the presence of copper ions. 


Note: The formation of the violet colour confirms that the proteins are present.


  1. Xanthoproteic Test

When proteins are treated with nitric acid, it gives an orange or yellow colour. The concentrated nitric acid gets used for the nitration process. When proteins are treated with nitric acid, it gives a yellow precipitate that later turns to orange colour when it is treated with an alkali.


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Note: The appearance of the yellow coloured solution confirms that proteins are present.


  1. Millon’s Test

The phenolic group of the tyrosine compound of proteins tends to react to the mercuric sulphate in the presence of sulphuric acid and sodium nitrate and gives a red colour. The Million’s test is done on the proteins that have phenolic amino acids. However, gelatin does not show this test. Initially, a white precipitate is formed when the proteins are treated with the Million’s reagent and they then turn into brick red in colour when boiled. This confirms that proteins are present.


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Note: When a brick red solution appears, it confirms the presence of the proteins.


  1. Ninhydrin Test

Proteins undergo a reaction with the pyridine solution of ninhydrin to give a coloured solution. This colour ranges from deep blue to violet-pink and even red in a few cases. The ninhydrin solution is prepared when 0.1g of ninhydrin is dissolved in 100ml distilled water. However, this ninhydrin is highly unstable and can only be kept for 2 days. The chemical reaction is as follows.


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Note: The appearance of the violet coloured solution confirms that the proteins are present.


Materials Required for the Protein Lab Test 

The list of materials are given below that are required for the protein lab test.

  1. Sodium hydroxide

  2. Pyridine solution

  3. Ninhydrin reagent

  4. Copper sulfate solution

  5. Nitric acid

  6. Sodium nitrite

  7. Sulfuric acid

  8. Mercuric sulfate

  9. Distilled water

  10. Dropper

  11. Test tubes

  12. Test tube holder

  13. Stirrer

  14. Water bath


Apparatus Setup of the Protein Total Spot Urine Test

Given below is the setup for the test to give low protein lab results.


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Procedure

The procedures of the laboratory test of proteins are as follows:.

  1. Biuret Test:

  • Take the sample given that is to be tested in a test tube.

  • Then add 2ml of the sodium hydroxide solution to the sample.

  • To this add about 5 to 6 drops of the copper sulfate solution.

  • If a bluish violet colour appears, it indicates the presence of the proteins.


  1. Xanthoproteic Test:

  • Take 2ml of the sample in a clean test tube.

  • Add a few drops of the concentrated sulfuric acid to it and then heat.

  • If a yellow precipitate is seen to be formed, it confirms the presence of the proteins.


  1. Millions Test:

  • Take 2ml of sample solution given in a test tube.

  • To it add about 2–3 drops of the Millon’s reagent and then shake well.

  • Observe any changes.

  • If a white precipitate is seen which then changes to brick red when heated, then it confirms the presence of the proteins.


  1. Ninhydrin Test:

  • Take the given sample solution that is to be tested in a test tube.

  • Add about 1–2ml of the ninhydrin solution to it.

  • Then boil the mixture and observe any changes.

  • If the solution turns blue in colour, then it confirms the presence of the proteins.


Observation and Inference from the Protein Laboratory Test


Biuret Test

The formation of violet colour confirms that the proteins are present.

Xanthoproteic Test

The yellow coloured solution that appears confirms the presence of the proteins.

Millions Test

The brick red colour solution confirms that the proteins are present.

Ninhydrin Test

The violet coloured solution confirms that the proteins are present.


Results and Discussions

The given sample contains _________ (proteins).


Precautions

  1. Handle the chemicals with care when performing the experiments and tests.

  2. Wear lab aprons and gloves while performing the experiments.

  3. Use only the droppers for taking the reagents from the bottles.


Viva Questions for the Total Protein Lab Test

1. What are proteins?

Answer: Proteins are made of smaller units known as amino acids which are attached in the form of longer chains to one another. Twenty different types of amino acids can be combined together to form a protein molecule.


2. What are the two different types of proteins?

Answer: The two different kinds of proteins are fibrous proteins and globular proteins.


3. What happens when a protein undergoes hydrolysis?

Answer: When proteins undergo the process of hydrolysis, they form α-amino acids.


4. How can amino acids together form a polypeptide?

Answer: Two amino acids get linked together when a water molecule is lost. The amino acids that are joined together by the peptide bonds tend to form a chain of polypeptides and every unit of the amino acids in the polypeptide is referred to as the residue. 


5. What do you mean by the monomer and polymer of the proteins?

Answer: A monomer is referred to as a molecule that forms bigger polymer molecules. It is known to be the building block of proteins. For example, amino acids are the building blocks of proteins. A polymer is known as a protein of the monomer series.


Conclusion

There are many important functions that are performed by proteins which are essential for all forms of life. They are important structural molecules. In this article, we learn all the necessary information of laboratory tests of proteins.


FAQs on Laboratory Test of Proteins

1. How will you identify a protein by laboratory test?

There are some chemical tests that are helpful in identifying the presence of protein in solutions along with other biological molecules, for example, milk. The presence of proteins in biological fluids is very important biologically. Hence, its presence in any solution can be easily identified by the result of different biological tests such as biuret test (result: formation of violet colouration), 


xanthoproteic test (result: the appearance of yellow colour solution), Million’s test (result: the appearance of brick red colour solution confirms the presence of proteins), and ninhydrin test (result: the appearance of violet colour solution).

2. What happens if our body is protein deficient?

Protein is very important for the growth of the human body. Protein deficiency is linked with malnutrition and inadequate protein intake. The deficiency of protein can cause serious health problems such as porous bones that are weak and thin, swelling (stomach bloating), stunt growth in children, fatty liver, increase the severity of infections and skin degeneration, etc. The low intake of protein may cause muscle wasting as well; it also increases the risk of bone fractures. The requirement of the amount of protein depends on the age of the person.

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