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Characterization of lipoxins by combined gas chromatography and electron‐capture negative ion chemical ionization mass spectrometry: Formation of lipoxin A 4 by stimulated human whole blood
Author(s) -
Brezinski Damian A.,
Serhan Charles N.
Publication year - 1991
Publication title -
biological mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1052-9306
DOI - 10.1002/bms.1200200202
Subject(s) - chemistry , trimethylsilyl , chromatography , chemical ionization , lipoxin , gas chromatography , mass spectrometry , whole blood , isotope dilution , ether , organic chemistry , biochemistry , ion , ionization , biology , receptor , immunology
The lipoxins are a recent addition to the family of bioactive products derived from arachidonic acid. Here, we have prepared pentafluorobenzyl ester, trimethylsilyl ether derivatives of lipoxin A 4 , lipoxin B 4 and pentadeuterolipoxin A 4 and have characterized these products by electron‐capture negative ion chemical ionization gas chromatography/mass spectrometry (NICI GC/MS). Lipoxin A 4 (5 S ,6 R ,15 S ‐trihydroxy‐7,9,13‐ trans ‐11‐ cis ‐eicosa‐tetra‐enoic acid; LXA 4 ) was quantified following extraction from whole blood by stable isotopic dilution utilizing deuterium‐labeled LXA 4 as internal standard and selected ion monitoring of the [M ‐ pentafluorobenzyl] anions. Studies with a second tritiated internal standard (e.g. [11,12‐ 3 H]LXA 4 ) also showed that the recovery of LXA 4 was > 80% following solid‐phase extraction from whole blood, and > 90% from isolated cells. In addition, neither isolated neutrophils nor platelets oxidatively metabolized [11,12‐ 3 H]LXA 4 when incubated in the presence or absence of stimuli. Whole blood incubated with either the ionophore of divalent cations (A23187), thrombin, or thrombin plus the chemotactic peptide formylmethionyl‐leucine‐phenylalanine generated both LXA 4 and thromboxane, which were quantified by stable isotope dilution. The ratio of thromboxane to LXA 4 formed by stimulated whole blood ranged from ∼2:1 to 20:1. These results indicate that the lipoxins display suitable characteristics as their respective pentafluorobenzyl ester, trimethylsilyl ether derivatives for quantification by electron‐capture NICI GC/MS. Moreover, they provide evidence that LXA 4 can be generated from endogenous sources in whole blood following exposure to physiologically relevant stimuli.