Isotope-dilution method for the determination of 1-vinyl-2-pyrrolidone-mercapturic acid as a potential human biomarker for 1-vinyl-2-pyrrolidone via online SPE ESI-LC/MS/MS in negative ionization mode

- The manuscript provided deals with a new online SPE ESI-LC/MS/MS method for a yet unknown 1-vinyl-2-pyrrolidone (VP) metabolite, that is 1-vinyl-2-pyrrolidone mercapturic acid (VPMA).
- Comparisons to structural similar substances and their metabolites and analytical methods were made.
- A fast and simple but specific and sensitive automated online SPE program using restricted access material (RAM) is presented. Mass spectrometric tuning data with according mass transitions of standard and isotope labeled internal standards are presented.
- Thorough method validation data is presented.
- Chromatograms of samples from exposure experiments with Sprague-Dawley rats are given for the proof of principle.

We established and validated a specific and sensitive analytical method for the determination of 1-vinyl-2-pyrrolidone (VP) as 1-vinyl-2-pyrrolidone-mercapturic acid (VPMA) in urine using an electrospray liquid chromatography tandem mass spectrometry (ESI-LC/MS/MS) column switching method. An online solid phase extraction (SPE) for sample cleanup was performed by column switching to a restricted access material and back-flushing to the analytical column. A Phenomenex Luna C8 column was used for sample separation (150 mm; ID 4,6 mm; 3 μm). D4-VPMA served as an isotope labeled internal standard and was detected in negative multiple-reaction monitoring (MRM) mode.The Limit of quantification (LOQ) for VPMA was 1.5 μg/L, the intra-day precision of three concentrations (2 μg/L, 75 μg/L and 400 μg/L) of spiked urine samples ranged from 2.7 to 7.3%, the inter-day precision from 3.4 to 14.4%. The accuracy ranged from 6.2 to 9.0%, for the intra-day experiments and from 0.3 to 6.9% for the inter-day experiments. The method was applied to urines of Sprague-Dawley rats exposed to VP as a proof of principle of VPMA as a potential biomarker.