Improved enzymatic assay for plasma hydroperoxides, inactivation of interfering enzymes

The involvement of free oxygen radicals in the pathology of certain diseases explains the growing interest in the assay of lipid peroxides. Assay of polyunsaturated fatty acid degradation products is currently performed by measuring the so-called thiobarbituric acid-reacting substances, of which malondialdehyde is the best known. The fact that this assay is controversial and that a second index of free radical attack would be useful to confirm the peroxidative process prompted us to undertake this work. We propose an improved enzymatic technique for assay of lipid hydroperoxides in biological fluids. Our aim has been to improve the technique already described by Heath and Tappel, which cannot be used as described by the authors. In fact, the presence of endogenous enzymes such as glutathione peroxidase and glutathione reductase in the sample interferes with the reaction and makes the results unreliable. Elimination of these endogenous enzymes by deproteinization before assay of the lipid hydroperoxides in the plasma gives a simple, reliable, and reproducible assay if the experimental procedures described in this work are followed.