IDENTIFICATION OF ARACHIDONATE EPOXIDES DIOLS BY CAPILLARY CHROMATOGRAPHY MASS-SPECTROMETRY

The identification of epoxide regioisomers of arachidonic acid (EETs) as methyl esters is difficult because they coelute during gas chromato graphy and possess similar mass spectra. In the present study EETs and their hydrolysis products, dihydroxyeicosatrienoic acids (DHETs), wer e analyzed as pentafluorobenzyl ester derivatives and their properties were compared to other esters. The four EET regioisomers were not res olved by gas chromatography as pentafluorobenzyl, trimethylsilyl, t-bu tyldimethylsilyl, or methyl esters. However, after being hydrolyzed to DHETs, three of the four regioisomers were resolved as (bis)-t-butyld imethylsilyl ether, pentafluorobenzyl esters. The fourth regioisomer ( 5,6-DHET) was resolved after being converted to a delta-lactone. Thus, the EETs could be resolved by capillary gas chromatography once conve rted to DHETs. Pentafluorobenzyl esters of both EET and DHETs (15-40 n g) provided diagnostic spectra when analyzed by electron ionization ma ss spectrometry. The mass spectral interpretations that indicated epox ide and diol positions were validated using synthesized EET/DHET [17,1 7,18,18-d(4), 5,6,8,9,11,12,14,15 d(B)] standards. Lesser amounts of D HETs (5-150 fg) also indicated molecular weights when analyzed in the negative-ion chemical-ionization mode. In summary, EETs in nanogram qu antities were identified as pentafluorobenzyl esters using electron io nization mass spectrometry. EETs in femtogram-to-picogram quantities w ere also identified after conversion to DHETs and analysis by gas chro matography-mass spectrometry in the negative ion-chemical ionization m ode.