Simultaneous quantification of S-adenosyl methionine and S-adenosyl homocysteine in human plasma by stable-isotope dilution ultra performance liquid chromatography tandem mass spectrometry.
S-adenosyl methionine (SAM) is an important methyl group donor that is formed from methionine. S-adenosyl homocysteine (SAH) is formed after demethylation of SAM and represents a potent inhibitor of many methyltransferases. We developed an improved stable-isotope dilution ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the simultaneous quantification of SAM and SAH in biological samples. The method comprises a phenylboronic acid-containing solid-phase extraction procedure, serving for binding and clean-up of SAM and SAH. After extraction, samples were separated and detected using either a HPLC SymmetryShield RP(18) or an Acquity UPLC BEH C(18) column with a HPLC-MS/MS or an UPLC-MS/MS system. The best results were obtained by Acquity UPLC BEH C(18) column. In plasma samples, the estimated intraassay coefficients of variation (CVs) for SAM and SAH were 3.3% and 3.9%, respectively, the interassay CVs were 10.1% for SAM and 8.3% for SAH. Mean recovery of SAM and SAH at two different concentrations was 100.0% for SAM and 101.7% for SAH. The quantification limits were 0.5 and 0.7nmol/L for SAM and SAH, respectively. In 31 plasma samples, the mean concentrations (SD) were 85.5 (11.1)nmol/L for SAM and 13.3 (5.0)nmol/L for SAH with a SAM/SAH ratio of 7.0 (1.8). The new UPLC-MS/MS method showed very high sensitivity and selectivity for SAM and SAH, low CVs and fast sample preparation (40 samples in 60min) and analysis time (3min). This new assay can be used for large-scale clinical studies.
Kirsch
Susanne H
SH
Saarland University Hospital, Department of Clinical Chemistry and Laboratory Medicine, Building 57, 66421 Homburg/Saar, Germany.
Knapp
Jean-Pierre
JP
Geisel
Jürgen
J
Herrmann
Wolfgang
W
Obeid
Rima
R
D000328:Adult; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D007553:Isotope Labeling; D008297:Male; D008875:Middle Aged; D012435:S-Adenosylhomocysteine / Q000097:blood*; D012436:S-Adenosylmethionine / Q000097:blood*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D055815:Young Adult
D012436:S-Adenosylmethionine; D012435:S-Adenosylhomocysteine