Qualitative Protein Analysis

Qualitative tests and the analyses of these are done to identify the structure and reaction of each protein to a particular test. There are general and specific tests to be able to identify the different types of proteins clearly and to classify them into groups. General tests include the Biuret and Ninhydrin while for the specific types of tests, these include the Xanthoproteic, Million-Nasse, Hopkins-Cole, Sakaguchi and Lead Acetate.

Biuret Test. The Biuret Test is positive for peptide bonds in the proteins. According to Koffuor (2012), the Biuret Test is used in the detection and estimation of proteins and peptides having more than two amino-acids. The reaction is characterized by the addition of copper sulfate to compounds with two peptide

The Ninhydrin Test is the general test used to identify both proteins and amino acids. It can be used qualitatively and quantitatively. For example, chromatographic visualization for the former and peptide sequencing for the latter. The colors produced are because of the amines reacting with the ninhydrin. Different amines produce different colors. For example, α-amino acids produce a blue-purple product while secondary amines like Proline produce a yellow-orange product (Hunt, n.d.). In the Ninhydrin Test, the electron deficient polar carbonyl carbon is attacked by the nucleophilic nitrogen on the amino acid. This combines the ninhydrin to the amino acid molecule temporarily. Until the carbon originally attacked is protonated and leaves in water form, the structure stays rearranged. When the nitrogen is double bonded to the originally attacked carbon, this creates a Schiff base. Once again, the molecule rearranges so the nitrogen would be double bonded to the adjacent carbon of the amino acid. The last rearrangement of the molecule produces a carbon dioxide gas. Any more rearranging produces ruhemann's

The Lead Acetate Test is a specific test in indicating the presence of sulfur in the protein chain. This test only shows a positive result in cysteine and cystine, the only types of protein containing sulfur. In the experiment, the reagent used was lead acetate (Pb(OAc)2) in NaOH. As explained in Milio and Loffredo (n.d.), boiling of cysteine and sodium hydroxide (NaOH) in a water bath converts the sulfur in the protein into sodium sulfate (NaS), causing the precipitation of lead from the solution as observed with the presence of black precipitate once the reaction occurred. The presence of the black precipitate indicates that sulfide and not sulfate, which gives off a brown color, was present in the reaction.

Precipitation reactions. Precipitation reactions involve the denaturation of proteins. This is so because the most common observation with protein denaturation is the coagulation or precipitation of the product. Denaturation is the disruption and/or the possible destruction of the secondary and tertiary structures in the protein. The sequence of amino acids or the primary structures remain the same as the denaturation reactions are not strong enough to break the peptide bonds therefore only disrupting the normal alpha-helix and beta sheets and disentangling them into random