IDENTIFICATION AND RELATIVE QUANTITATION OF F-2-ISOPROSTANE REGIOISOMERS FORMED IN-VIVO IN THE RAT

F-2-isoprostanes are a complex mixture of isomers formed in four regio isomeric family types by free radical-initiated oxidation of arachidon ic acid present in membrane phospholipids. F-2-isoprostanes isolated f rom the livers of rats treated with carbon tetrachloride were separate d by initial reverse phase HPLC and detected using electrospray ioniza tion mass spectrometry with the characteristic loss of 44 u (C2H4O) fr om the common 1,3-diol cyclopropane ring found in these eicosanoids. C ollision induced decomposition of the carboxylate anions from the sepa rated F-2-isoprostanes formed abundant ions characteristic for regiois omers of Type I (m/z 115), Type LII (m/z 127), and Type IV ( m/z 193), which made possible characterization of these three family subtypes b y LC/MS/MS. Capillary GC/MS was employed to further identify the F-2-i soprostane regioisomers using electron ionization mass spectrometry an d to obtain characteristic mass spectra of the pentafluorobenzyl ester trimethylsilyl ether derivatives. Quantitation of the F-2-isoprostane s separated by both reverse-phase HPLC and capillary GC/MS was carried out using negative ion chemical ionization mass spectrometry. The mos t abundant isomers identified were Type I and IV regioisomers constitu ting 33 and 25% of the total products, respectively. As expected, the Type II and III regioisomer products were of less abundance. Over 45 F -2-isoprostanes could be separated in this complex mixture, suggesting random production of each regioisomeric subtype in this in vivo model . (C) 1997 Elsevier Science Inc.