Schiff’s test

Schiff’s test

A biochemical test to detect cholesterol in solution.


When cholesterol solution is mixed with concentrated sulphuric acid without disturbing the layers, a red ring is formed at the interface of the two layers. The red colour is due to the formation of Dicholestadiene, a condensation product between two cholesterol molecules with removal of one molecule of water.



  • Cholesterol solution
  • Chloroform
  • 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.


  • Take 1 ml of cholesterol solution contained in a dry test tube.
  • Add 0.5 ml of concentrated sulphuric acid through the wall without shaking.
  • Stand it for sometime, see the development of red ring at the junction of two layers.



  • Appearance of red ring at the junction of two layers indicates presence of cholesterol in solution.


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