Solid- and liquid-phase extraction for the gas chromatographic–tandem mass spectrometric quantification of 2,3-dinor-thromboxane B 2 and 2,3-dinor-6-oxo-prostaglandin F 1α in human urine

Abstract

Whole body synthesis of thromboxane A 2 is best assessed by quantifying non-invasively its major urinary metabolite, i.e., 2,3-dinor-thromboxane B 2 (2,3-dn-TxB 2), by gas chromatography–mass spectrometry (GC–MS) or GC–tandem MS. Methods based on these techniques usually require a series of extraction and purification procedures including solid-phase extraction (SPE) and thin-layer chromatography (TLC) or liquid chromatographic separation of authentic or derivatized 2,3-dn-TxB 2. Taking advantage of the inherent accuracy of GC–tandem MS and the high selectivity of the extraction of methoximated 2,3-dn-TxB 2 on phenylboronic acid SPE cartridges we developed a method that involves only SPE steps prior to quantification by GC–tandem MS. The method was validated by performing in parallel an additional TLC step. Method mean accuracy and precision were of the order of 103% and 95%, respectively. The method allows furthermore co-processing of the same urine sample to quantify accurately and rapidly the major urinary metabolite of prostacyclin, i.e., 2,3-dn-6-oxo-prostaglandin (PG) F 1α, by GC–tandem MS. The limit of detection of the method was below each 5 pg of 2,3-dn-TxB 2 and 2,3-dn-6-oxo-PGF 1α per 5 ml of urine. Our study suggests that dinor metabolites of isothromboxanes and isoprostacyclins are not abundantly present in human urine.