Quantitative Analysis of 5-Oxo-6,8,11,14-eicosatetraenoic Acid by Electrospray Mass Spectrometry Using a Deuterium-Labeled Internal Standard

✍ Scribed by William S. Powell; Daniel Boismenu; Subhash P. Khanapure; Joshua Rokach

• Publisher: Elsevier Science
• Year: 2001
• Tongue: English
• Weight: 66 KB
• Volume: 295
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.2001.5206

✦ Synopsis

5-Oxo-6,8,11,14-eicosatetraenoic acid (5-oxo-ETE), a metabolite of arachidonic acid formed by the 5-lipoxygenase pathway, is a potent eosinophil chemoattractant that may be an important mediator in asthma. To further investigate the physiological and pathological roles of 5-oxo-ETE we have developed a mass spectrometric assay employing a tetradeuterated analog ( 5oxo-[11,12,14,15-2 H]ETE) as an internal standard. Collision-induced dissociation of the quasimolecular anion of 5-oxo-[11,12,14,15-2 H]ETE (m/z 321) resulted in the formation of a major ion at m/z 207 that retained all four deuterium atoms. Measurement of the ratio of ions at m/z 203 (endogenous 5-oxo-ETE) and m/z 207 permitted quantitation of this compound by liquid chromatography-mass spectrometry-mass spectrometry using multiple reaction monitoring. The resulting assay was highly sensitive (<20 pg/sample) and selective, enabling detection of the amount of 5-oxo-ETE produced by as few as 10,000 neutrophils. This assay should permit measurement of 5-oxo-ETE in biological fluids, enabling evaluation of its role in asthma and other inflammatory diseases.