We studied the kinetics of the reaction between cholesterol and sulfuric acid in acetic acid-acetic anhydride medium. Results have been used to establish near-optimal conditions for the fast kinetic determination of cholesterol in serum. The reaction rate measured during the first 20 s of the reaction is proportional to cholesterol concentration. There is good agreement (<2% deviation) between pseudo-first-order rate constants for cholesterol standards and sera. Recoveries of standard cholesterol added to sera range from 99% to 104% (average, 100.7%). Values for serum cholesterol by this kinetic determination tend to be somewhat lower than equilibrium values reported by local hospital laboratories. Whether bilirubin interferes depends on reaction conditions; under optimal conditions, each milligram of bilirubin is kinetically equivalent to about 1 mg of apparent cholesterol. Kinetic data are included to show how results are degraded by operating under nonoptimal conditions, and considerations involved in changing experimental conditions are discussed.
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