Solubility Tests of Proteins Principle, Procedure, Result, Application

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Solubility Tests of Proteins

Solubility Tests is a type of biochemical test which is performed to check the ability of compounds to dissolve within a liquid solvent.

  • It is an essential test which can easily determine the size and polarity of unknown compounds and as well as the presence of acidic and basic functional groups.
  • The Solubility tests for proteins and amino acids are considered as the qualitative test which can detect the presence and absence of different proteins.
  • By detecting the solubility of proteins we can gather information regarding their functionality.
  • The protein solubility in aqueous buffers depends on the distribution of hydrophilic (having a strong affinity for water) and hydrophobic (lacking affinity for water) amino acid residues on the surface of proteins.
  • Hydrophobic residues are predominantly found in the globular protein core, but some exist in patches on the surface.
  • Proteins that have high hydrophobic amino acid content on the surface have low solubility in aqueous solvents.
  • Charged and polar surface residues interact with ionic groups in the solvent and increase solubility.
  • Solvent media has a great effect on protein solubility. These effects are: Ionic Strength, pH, Temperature, Dielectric Constant

Aim of Solubility Tests of Proteins

  • To detect the solubility of proteins in different solvents.
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Principle of Solubility Tests of Proteins

The nitrogen present within the protein sample is proportional to the solubility of proteins, which remains in a soluble state under specific conditions. The amino acids of proteins are soluble in water while they are insoluble in non-polar organic solvents. The amino and carboxyl groups of amino acids helps the amino acids and proteins to accept and donate protons to the aqueous solution.

The protein solubility within a solvent is affected by different factors such as the molecular size, pH of the medium, hydration of the proteins, and the salting-in process. The addition of salts during the salting-in technique, generates an interaction between the surface ionic charges on the proteins and the electrolyte, which prevents the aggregation of protein particles. Depending on all of these factors, the solubility of the protein in different solvents might differ.

Requirements for Solubility Tests of Proteins

  • Reagent: Sample solution, Solvents (HCl, NaOH, Chloroform, Water)
  • Materials: Test tubes, Test tube stand, Dropper.

Solubility Tests of Proteins Procedure

  1. A small amount of different solvents are taken in different test tubes.
  2. Add a few drops of the protein sample solution and mix properly.
  3. Allow the test tubes to stand for about 5 minutes, and then the observations are made.

Result of Solubility Tests of Proteins

  • Positive Test: The appearance of a clear solution with no precipitation of cloudiness.
  • Negative Test:  The appearance of a cloudy solution with precipitation.

Application of Solubility Tests of Proteins

  • This test helps us to understand the pattern of solubility of proteins which is then helps in the detection and differentiation of such proteins.
  • The solubility of proteins gives insight into the solubility of proteins during the development and testing of new protein composition.
  • This test allows the selection of proteins for their application in liquid foods and beverages.
  • It also provides information about the amino acid composition and molecular weight.

Limitations of Solubility Tests of Proteins

  • Carefully carried out the observation step of solubility tests, there is a chance of false-negative results might be prevalent.
  • Allow the test tubes to stand for some time to provide some time for solubility.


  • Zayas J.F. (1997) Solubility of Proteins. In: Functionality of Proteins in Food. Springer, Berlin, Heidelberg.
  • D (2012). Biochemistry. Fourteenth Edition. Academic Publishers. Kolkata.

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