Libermaann-Burchard Test  Or Acetic Anhydride Test

Libermaann-Burchard Test  Or Acetic Anhydride Test

(For the qualitative detection of cholesterol)


The Liebermann–Burchard  test is used for the detection of cholesterol. It is also known as acetic anhydride test. To a solution of cholesterol in chloroform, when a few drops of acetic anhydride is added, mixed well, and a few drops of concentrated sulphuric acid added from side of the tube, kept in dark. A red rose colour develops which changes rapidly to deep green. One molecule of water is removed from cholesterol, and it is oxidized giving rise to green fluorescent colour.


  • Cholesterol solution
  • Chloroform
  • Acetic anhydride
  • Concentrated sulphuric acid

Sample preparation:

  • Separate the egg yolk (yellow part) of a boiled egg and grind it with about 50 ml of chloroform in a mortar.
  • Filter the suspension through a filter paper.
  • The filtrate is the cholesterol solution. The egg yolk is the rich source of cholesterol.


  • Dispense about 2 ml of cholesterol solution in dry test tube.
  • Add 10 drops of acetic anhydride and mix well.
  • Then, add 2 drops of conc. sulphuric acid from side of tube wall.
  • Keep it in dark for about 5 mins, and observe the appearance of green colour.



Appearance of green colour indicates presence of cholesterol in solution.


Sulphuric acid and acetic anhydride are corrosive and hazardous


Leave a Reply

Your email address will not be published. Required fields are marked *