Assay of urinary 2,3-dinor-6-oxo prostaglandin F1α by gas chromatography—tandem mass spectrometry

Abstract

Prostacyclin (PGI2), an important determinant of cardiovascular biology, is produced from arachidonic acid by endothelial cells. Measurement of its stable urinary metabolite, 2,3-dinor-6-oxo prostaglandin F1 alpha (PGI2-M), is the approach of choice to assess variations of the endogenous synthesis of PGI2 that occur in response to dietary, pharmacological and pathological alterations. We developed a relatively simple stable isotope dilution assay for PGI2-M which involves solid-phase extraction of 10 ml of urine with Chem Elut disposable columns, water/solvent partitioning from basic and acidic environments with ethyl acetate and methylene chloride, derivatization to 1-pentafluorobenzyl ester followed by TLC, methoximation and trimethylsilylation. Quantification was achieved, for the first time for PGI2-M, by capillary GC-electron capture negative ion MS-MS with a triple quadrupole mass spectrometer operated in the negative ion detection mode with methane as moderating gas. The mean inter-assay R.S.D., determined on 12 different urine samples, was 5.1 (range 0.4% to 10.5%). The excretion of PGI2-M in 34 healthy male subjects (age 26 to 57) was 156.2 +/- 65.2 (mean +/- S.D.) ng/24 h.