Isoprostanes assays

D, Dworski R, Kanai K, Taber D, Moore K, Oates JA, 23. Roberts LJ. Quantification of the major urinary metaboUte of 15-F2t-isoprostane (8-iso-PGF2alpha) by a stable isotope dilution mass spectrometric assay. Anal. Biochem. 1999 269 326. 

The assays most widely employed are the measurement of thiobarbituric acid-reactive species (TBARS) and the formation of conjugated dienes, markers of lipid peroxidation [31-33] the determination of advanced oxidation protein products (AOPP), a marker of protein oxidation, and of advanced glycation end-products (AGE) [34-37] the measurement of erythrocyte antioxidant potential [38]. Of particular importance is the isoprostanes determination, since these compounds are formed by the free radical catalysed peroxidation of arachidonic acid, which is independent of the cyclooxygenase enzyme, giving rise to stable compounds, measurable in urine [39]. As recently assessed in a Workshop on markers of oxidative damage and antioxidant protection [40], currently available methods for the determination of antioxidant and redox status are not yet generally suitable for routine clinical applications, essentially for the lack of standardized tests. 

Morales CR, Terry ES, Zackert WE, Montine TJ and Morrow JD, Improved assay for the quantification of the major urinary metabolite of the isoprostane 15-F(2t)-Isoprostane (8-iso-PGF(2alpha)) by a stable isotope dilution mass spectrometric assay. Clin Chim Acta 314(1-2) 93-9. 2001. 

One approach to determination of whole-body lipid peroxidation has been measurement of exhaled hydrocarbons by GLC, especially ethane. Hydrocarbon gases are, however, minor end-products of peroxidation and their formation depends on the decomposition of peroxide. Recent studies have demonstrated that isoprostane is a good biomarker of lipid peroxidation in the human body. Isoprostanes are specific products arising from the peroxidation of unsaturated fatty acid residues in lipids and detection of them and their metabolites in urine is a useful assay of whole-body lipid peroxidation. Isoprostanes can be accurately and sensitively measured by mass spec-trometric techniques. 

These measurements include the F2 isoprostanes—the oxidation products of arachi-donic acid—and methods require mass spectrometry or immunoassay. Other measures of lipid peroxidation include hydroperoxides by luminometry or colorimetric assays, or a third colorimetric measurement of aldehyde products (TEAR) with thiobarbituric acid where malondialdehyde is measured malondialdehyde is an end-product derived from the breakdown of polyunsaturated fatty acids and esters. 

Most of the assays of isoprostanes to date have focused on assessing 8-iso-PGF levels in body fluids, since this is a major product of the total peroxidation process in vivo. When measuring the urinary metabolites of S-iso-PGF, the correct choice of the appropriate metabolite is important since the metabolic profile and appearance of the different metabolites vary between species (Roberts et al, 1996 Basu, 1998c Chiabrando et al, 1999). The tetranor metabolite of 8-iso-PGFj is the major urinary product in the rabbit, whereas the dinor metabolite is the dominant product in humans. 

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