Xanthoproteic Test for Tyrosine and Tryptophan

Reaction of concentrated nitric acid with some substituted aromatic rings gives a yellow color (Xanthos = yellow in Greek).

Procedure

    Set up test tubes as follows: In one tube put 2 drops of tryptophan solution, in another 2 drops of unknown compound, in another 2 drops of glycine solution, and in another about 5 mg tryosine powder. Then with great care, add 1.0 mL concentrated nitric acid (HNO₃) to each test tube. Hold the tubes so that they do not point at you or anyone else.  Heat the tubes gently in a water bath until they boil. Cool the tubes slowly and add the 4% NaOH, drop by drop, until the solutions are alkaline. 

Did any color change take place before the alkali was added? What was the final color of each of the solutions?

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Principles of Gel Electrophoresis

Electrophoresis: How to Read Results

Protein Purification with Dialysis Tubing

Protein Quantification (Bradford Assay)

Bradford assay is a rapid and accurate method to determine the concentration of protein. The assay is based on the observation that the absorbance maximum for an acidic solution of Coomassie Brilliant Blue G-250 shifts from 465 nm to 595 nm when binding to protein occurs. Both hydrophobic and ionic interactions stabilize the anionic form of the dye, causing a visible color change. Within the linear range of the assay (~5-25 mcg/ml), the more protein present, the more Coomassie binds.