Quantification of citrulline by parallel fragmentation monitoring — A novel method using graphitized carbon nanoparticles and MALDI-TOF/TOF mass spectrometry

BackgroundSelected reaction monitoring (SRM) is a reliable mass spectrometry (MS)-based technique for quantification of small molecules. However, it is not applicable to matrix-assisted laser desorption/ionization time-of-flight/time-of-flight tandem MS (MALDI-TOF/TOF MS) instruments. This work presents a novel comparable MALDI-TOF/TOF MS technique, “Parallel Fragmentation Monitoring” (PFM), for high-throughput quantification of citrulline.MethodCalibrator/sample solutions were spiked with internal standard that was a stable isotopic analog with 1 mass unit heavier than citrulline. Both citrulline and internal standard were isolated and fragmented in parallel by MALDI-TOF/TOF MS in the presence of graphitized carbon nanoparticles as matrix. The ratio of the peak intensities of the selected fragment of citrulline to that of internal standard was used to calibrate/calculate the concentrations of citrulline in samples.ResultsLinear calibration curves were obtained in the range of 10–250 μmol/l citrulline with correlation coefficients ≥ 0.997. Stored calibration curve and batch-specific calibration curve produced highly similar measurement values. Within- and between-day CVs were 3.1–8.7% and 3.5–10.6%, respectively, illustrating the reliability and robustness of PFM.ConclusionUsing citrulline for proof-of-concept, we have developed the PFM technique with tremendous potential for high-throughput quantification of amino acids and other small molecules.

► We present a new method for quantification of small molecules by MALDI-TOF/TOF MS.
► Mesoporous graphitized carbon nanoparticles were used as matrix to assist desorption/ionization.
► Precursor ions of citrulline and stable isotopic internal standard were isolated and fragmented in parallel.
► Peak intensity ratio of selected fragment of citrulline to that of internal standard was used in calibration/measurement.
► By using citrulline, this PFM method was shown to be accurate, precise and robust.