Define Principle for Estimation of cholesterol - Lipids?

Zlatkis and associates have evolved a method of quantitative determination of cholesterol based on a reagent containing ferric chloride, glacial acetic acid and concentrated sulphuric acid. Cholesterol gives a purple colour with this reagent. The colour development is due to a dehydration reaction in the cholesterol molecule to form 3, 5 cholestadiene which is oxidized by H₂SO₄ and polymerises to form a diamer or trimer. The cholastadiene and its polymers react with H₂SO₄ to form mono and disulfonic acids which are highly coloured. In the presence of added metal ions like ferric, the disulphonic acids are preferentially formed. The detailed step-by-step procedure for cholesterol estimation is included in Experiment 3 later in this practical. Study the procedure carefully before conducting the experiment. With this basic understanding about iodine number, saponification technique and methods for cholesterol estimation, we are well equipped to carry out the experiments given in this practical. There are three experiments in this practical.