Simultaneous determination of the primary prostanoids prostaglandin E2, prostaglandin F2 alpha and 6-oxoprostaglandin F1 alpha by immunoaffinity chromatography in combination with negative ion chemical ionization gas chromatography-tandem mass spectrometry.

The simultaneous determination of prostaglandin E2 (PGE2), prostaglandin F2 alpha (PGF2 alpha) and 6-oxoprostaglandin F1 alpha (6-oxo-PGF1 alpha) in urine using immunoaffinity chromatography in combination with negative ion chemical ionization gas chromatography-tandem mass spectrometry (NICI-GC-MS-MS) is described. Monoclonal antibodies against PGE2 (100% cross-reactivity with 6-oxo-PGF1 alpha) and PGF2 alpha were both coupled to a derivatized agarose matrix. After extraction with a C18 cartridge the sample was applied to the immunoaffinity column. The prostaglandins with eluted with acetone-water and the methoxime-pentafluorobenzyl-trimethylsilyl (MO-PFB-TMS) derivatives (PGE2 and 6-oxo-PGF1 alpha) and the PFB-TMS derivative (PGF2 alpha) were quantified by GC-MS-MS. For reproducibility experiments, spiked urine samples were analysed several times. The correlation coefficients were 0.997 (6-oxo-PGF1 alpha) and 0.999 (PGE2 and PGF2 alpha) and the slopes were 0.99 and 1.03, respectively. The inter-assay coefficient of variation ranged from 8.6 to 9.5% for the unspiked urine samples and from 2.0 to 5.2% for the spiked samples. This method offers several advantages, e.g., high specificity and sensitivity, good reproducibility and an increase in sample throughput.