Biuret reagent
The Biuret reagent is of potassium hydroxide (KOH) and copper (II) sulfate (CuSO4), together with potassium sodium tartrate (KNaC4H4O6·4H2O). The blue reagent turns violet in the presence of proteins, and changes to pink when combined with short-chain polypeptides. The potassium hydroxide does not participate in the reaction at all, but is merely there to provide an alkaline medium so that the reaction can take place.
The reagent is commonly used in a biuret protein assay, a colorimetric assay used to determine protein concentration--such as UV-VIS at wavelength 560nm (to detect the Cu2+ ion).
The Biuret Test
- Main Article: Biuret test
The principle underlying the test can easily be demonstrated with the chemical compound biuret which, just as proteins, is able to complex copper (II) ions. The biuret assay does not in fact use biuret, but is so named as it detects the peptide bond between the urea molecules or between amino acids.
Not all biuret tests actually require the Biuret reagent. Rather, the term "biuret test" is a generic term for the testing of proteins by using copper (II) sulfate solution in an alkaline environment. Thus, the process of testing for proteins in a solution by first adding a small amount of sodium hydroxide, and then adding copper (II) sulfate drop by drop, is also known as a biuret test.