Submitted on July 21, 2006
Revised on August 25, 2006
Accepted on August 29, 2006
Oxidized derivatives of 
-3 fatty acids; Identification of iPF3
-VI in human urine
John A. Lawson, Seongjin Kim, William S. Powell, Garret A. FitzGerald, and Joshua Rokach
Claude Pepper Inst. & Dept. of Chemistry, Florida Institute of Technology, Melbourne, FL 32901
Corresponding Author: jrokach{at}fit.edu
Isoprostanes (iPs) are prostaglandin-like molecules derived from autoxidation of polyunsaturated fatty acids (PUFAs). Urinary iP levels have been used as indices of in vivo lipid peroxidation. Thus far, it has only been possible to measure iPs derived from arachidonic acid (AA) in urine, as levels of iPs/neuroprostanes (nPs) derived from 
3-PUFAs have been found to be below detection limits of available assays. Because of the interest in 3-PUFA dietary supplementation, we developed specific methods to measure nPF4a-VI and iPF3a-VI (derived from DHA and EPA) using a combination of chemical synthesis, gas chromatography (GC)/MS and liquid chromatography (LC)/MS/MS. Although nPF4a-VI was below the detection limit of the assay, we conclusively identified iPF3a-VI in human urine by GC/MS and LC/MS/MS, the mean levels in 26 subjects being ~300 pg/mg creatinine. Our failure to detect nPF4a-VI may have been due to its rapid metabolism by ß-oxidation to iPF3a-VI, which we showed to occur in rat liver homogenates. In contrast, iPF3a-VI is highly resistant to ß-oxidation in vitro. Thus iPF3a-VI can be formed by two mechanisms: (i) direct autoxidation of EPA, and (ii) ß-oxidation of nPF4a-VI, formed by autoxidation of DHA. This iP may therefore serve as an excellent marker for the combined in vivo peroxidation of EPA and DHA in vivo.
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