Biuret’s test is used to indicate the presence of peptide bond in a substance being tested. The presence of the peptide bond indicates the presence of protein, as the amino acid binds each other by peptide bonds to form protein. Peptide bond is formed when carbon atom of the carboxyl group of a molecule bonded to a nitrogen atom of the amine group of other molecules. These reactions release water molecules so called condensation reaction.
The picture above shows the presence of two amino acid molecules binded by peptide bond to form protein. The peptide bond will react with biuret reagent produces a color change. Positive reaction is indicated by the appearance of purple or pink due to the reaction of Cu ++ of biuret reagent, NH of a peptide bond, and O of the water. The longer the bond peptide (more amino acids bonded) will bring up the color purple, the shorter the bond peptide (less amino acids bonded) will bring up the color pink.
Materials and reagents:
- NaOH 10%, 0.1% CuSO4
- The materials will be tested in solution form.
- Put 2 ml of the materials to be tested in a test tube.
- Enter 2 ml of NaOH 10% in the same tube.
- Drops with 1-10 drops of CuSO4 0.1% by a pipette.
- Observe the color change.
Biuret’s test is used for protein testing in general. Biuret’s will produce negative result on free amino acids because it doesn’t have a peptide bond.
Positive result by biuret’s test: boiled egg white, pasta meat, milk
Negative result by biuret’s test: egg yolk, flour, sugar