Abstract
Background: Available methods for the determination of nanomolar concentrations of S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) in plasma and cerebrospinal fluid (CSF) are time-consuming. We wished to develop a method for their rapid and simultaneous measurement.
Methods: We used tandem mass spectrometry (MS/MS) for the simultaneous determination of SAM and SAH, with stable-isotope-labeled internal standards. The 13C5-SAH internal standard was enzymatically prepared using SAH-hydrolase and [13C5]adenosine. The method comprises a weak anion-exchange solid-phase extraction procedure serving as clean-up step for the deproteinized plasma and CSF samples. After clean-up, samples were injected on a C18 HPLC column, which was connected directly to the tandem mass spectrometer, operating in MS/MS mode.
Results: In plasma samples, the intraassay CVs for SAM and SAH were 4.2% and 4.0%, respectively, and the interassay CVs were 7.6% and 5.9%, respectively. In CSF, the intraassay CVs for SAM and SAH were 6.8% and 6.9%, respectively, and the interassay CVs were 4.2% and 5.5%, respectively. Mean recovery of SAM and SAH for both matrices at two concentrations was 93%. Detection limits for SAM and SAH in samples were 7.5 and 2.5 nmol/L, respectively. Concentrations of SAM and SAH in plasma from healthy subjects were within the previously reported ranges. In 10 CSF samples, the mean concentrations (range) were 248 (137–385) nmol/L for SAM and 11.3 (8.9–14.1) nmol/L for SAH.
Conclusions: SAM and SAH can be analyzed by MS/MS, taking optimal advantage of the speed and high sensitivity and specificity of this relatively new analytical technique.
Simultaneous monitoring of monoclonal antibody variants by strong cation-exchange chromatography hyphenated to mass spectrometry to assess biosimilar-ity of rituximab-based biotherapeutics
