Quantitative Analysis of Brain Lipid Oxidation

Brain samples were prepared and analyzed for esterified 8-isoprostanes (8-IsoP) and prostaglandins (PGF2α) using gas chromatography-negative ion chemical ionization MS as described previously [19 (link)]. Separations were carried out on a DB-wax column (0.25 mm inner diameter × 15 m length × 0.25-μm film thickness). The oven temperature was programmed from 150 to 300 ºC held for 11 min. Samples were injected with a 1µL injection volume. Analyte peaks were obtained from ion chromatograms. [²H₄]-15-F2t-IsoP was used as internal standard (Cayman Chemical, Ann Arbor, MI USA). The major ion generated in the negative ion chemical ionization (NICI) mass spectrum of the pentafluorobenzyl (PFB) ester, trimethylsilyl ether (TMS) derivative of F2-IsoPs, was the m/z 569 carboxylate anion [M-181 (M-CH₂C₆F₅)]. The major ion generated in the NICI mass spectrum of the PFB ester, TMS ether derivative of F4-NPs, was the corresponding m/z 593 carboxylate anion. The ion generated by the [²H₄]-15-F2t-IsoP internal standard was m/z 573.