Biuret test is used for detecting compounds with peptide bonds. A biuret reagent may be used to test the aqueous sample. This blue reagent is made by combining sodium hydroxide and copper sulfate solutions. A few drops of this reagent will turn the aqueous sample containing compounds with peptide bonds from pale to intense violet colour. The violet colour intensity depends on the number of peptide bonds in the sample.
The biuret test is also used to detect proteins. That is because proteins are made up of polypeptides, which in turn, are made of amino acids joined by peptide bonds. The longer the polypeptide chain is, the more peptide bonds there are, and therefore, the more intense the violet colour will be when biuret test is applied. It also follows that a pale violet or pinkish colour indicates shorter polypeptide chains or fewer peptide bonds. A negative result (lack of violet colour formation) may mean lack of protein, or the presence of free amino acids (without peptide bonds).
The test, however, gives positive result to any compound containing two carbonyl groups attached to a nitrogen or carbon atom. Thus, it may not be completely protein-specific. Performing other protein tests may be necessary.