The development of a simple and highly sensitive assay for measuring lipid oxidation : an honors thesis (HONRS 499)
Oxidation is an essential process that is carried out in all cells. Free radicals are a byproduct of this process. Lipid oxidation, caused by damage by non-intercepted free radicals, is the cause of many degenerative diseases. Currently, all existing methods for measuring lipid oxidation have complications. These shortcomings, including lengthy procedures, slow reactions, strict experimental conditions, and low sensitivity, make the development of a new assay desirable. A new method was developed that overcame all of these shortcomings. Tertiary butyl hydroperoxide (t-BuOOH) was used to simulate an oxidized membrane. Diphenyl-lpyrenylphosphine (DPPP) was used to measure the oxidation by reacting with t-BuOOH and becoming diphenyl-l-pyrenylphosphine oxide (DPPPO), which fluoresces. This fluorescence was measured by using a High Performance Liquid Chromatography instrument for Flow Injection Analysis. An Fe3+/pyridine complex was used as a catalyst. During the process of developing this method, new complications arose, such as signal depression and reaction with molecular oxygen, but they were overcome. In the end, a highly sensitive and simple assay for measuring lipid oxidation had been developed.