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• CRPA (color reactions of proteins and amino acids)


  Used to detect linkage types, presence of amino acids, certain groups present, type of bonds
• Proteins


  Polymers of amino acids
• Ninhydrin reaction


  1. Also called triketohydrindenehydrate

  2. Powerful oxidizing agent

  3. Reacts with alpha-amino acids and proteins

  4. Intense blue or purple-colored complex

  5. Most general and one of the sensitive reactions known for the qualitative detection of proteins and their hydrolytic products
• Proline and hydroxyproline


  Give yellow color
• Free carbonyl group adjacent to the amino group


  CO2 is released
• Ninhydrin reaction is also given to ammonia, ammonium salts, primary amines, and amides
• Biuret reaction


  • Biuret is from heating urea at 180C
  • NH3OH (ammonium hydroxide) dissolves CuOH2 and form dark blue CuNH4++ ion
• Xanthoproteic reaction


  • Involves nitration of the benzene nucleus
  • Formation of yellow to orange color, makes the resulting compound alkaline
  • Positive reaction only to tryptophan and tyrosine
  • Tryptophan gives more of a red complex than tyrosine, partial oxidation of the substance by nitric acid
  • Phenylalanine also give off a positive reaction: after only once the PHE is nitrated by nitric acid and was added by a small amount of concentrated sulfuric acid
• Millon reaction


  • Monohydroxybenzene derivatives: tyrosine and phenol: positive reaction: faint pink to pure dark red
  • Dihydroxybenzene derivatives: resorcin and some indole derivate: yellowish to brownish red
  • Tyrosine is the monophenol derivative found in proteins
• Reduced sulfur test


  • Organic derivative of H2S is treated with a strong NaOH solution, it'll hydrolyze
  • Cysteine and cystine only, no methionine
• Sakaguchi reaction


  • Substances that contain guanidine group reacts with alpha-naphthol + oxidizing agent = orange or red color
  • Arginine gives a positive reaction