Perform the colour reactions on the given protein solution and report the amino acids present.
How to lay out the answer. Follow the frame, then write it in your own words.
1 · Aim. To perform the colour reactions of proteins on the given solution and identify the groups and amino acids present.
2 · Principle.
Biuret: Cu2+ and two or more peptide bonds in alkali give violet.
Ninhydrin: free α-amino groups give Ruhemann's purple.
Xanthoproteic and Millon's: aromatic rings and the phenol of tyrosine.
Hopkins-Cole: indole of tryptophan; Sakaguchi: guanidino group of arginine; lead acetate: sulphur of cysteine and cystine; Pauly's: histidine and tyrosine.
3 · Requirements. Egg albumin solution; 10% NaOH, 1% CuSO4, ninhydrin, conc. HNO3, Millon's reagent, Hopkins-Cole reagent, conc. H2SO4, α-naphthol, hypobromite, urea, 40% NaOH, lead acetate, Pauly's reagents; test tubes, water bath, pipettes with a bulb.
4 · Procedure.
2 mL protein for each test with the reagent volumes in your manual.
Heat where the test needs it (ninhydrin, xanthoproteic, Millon's, lead acetate).
Layer acid down the side for Hopkins-Cole.
5 · Observation. [Table: Test | Observation | Inference for all eight tests]
6 · Result. The given solution is a protein containing tyrosine, tryptophan, arginine, cysteine or cystine, and histidine.
7 · Precautions. Gloves and goggles; fume hood for concentrated acids; few drops of Millon's reagent; copper sulphate drop by drop; never pipette by mouth.
Scoring tip: Write the amino acid in the inference column for every test. 'Positive' alone earns little; 'brick red: tyrosine present' earns the mark.
