MCB Practical 2
Isolation and Characterisation of Caveolar Lipid Domains
Cholesterol Determination
Hydrogen peroxide is produced stoichiometrically to the amount
of cholesterol in the sample. In turn, this hydrogen peroxide reacts with
4-aminoantipyrine to produce a quinoneimine dye ( due to the increased
resonance path for electrons, this compound absorbs in the visible spectrum
-500nm). Prior to these two coupled reactions free cholesterol is released
from esters so that the A ring hydroxyl group is free for oxidation ( in
serum, about 60-70% of cholesterol is esterified).
Allain CC, Poon LS, Chan CSG, Richmond W, Fu PC " Enzymatic Determination
of Total Serum Cholesterol" Clin. Chem. 20 470-475 (1974)