Evaluation of multiple reaction monitoring cubed for the analysis of tachykinin related peptides in rat spinal cord using a hybrid triple quadrupole–linear ion trap mass spectrometer

摘要

Targeted peptide methods generally use HPLC-MS/MRM approaches. Although dependent onthe instrumental resolution, interferences may occur while performing analysis of complexbiological matrices. HPLC-MS/MRM3 is a technique, which provides a significantly betterselectivity, compared with HPLC-MS/MRM assay. HPLC-MS/MRM3 allows the detection andquantitation by enriching standard MRM with secondary product ions that are generated withinthe linear ion trap. Substance P (SP) and neurokinin A (NKA) are tachykinin peptides playing acentral role in pain transmission. The objective of this study was to verify whether HPLC-HPLCMS/MRM3 could provide significant advantages over a more traditional HPLC-MS/MRMassay for the quantification of SP and NKA in rat spinal cord. The results suggest thatreconstructed MRM3 chromatograms display significant improvements with the nearly completeelimination of interfering peaks but the sensitivity (i.e. signal-to-noise ratio) was severelyreduced. The precision (%CV) observed was between 3.5% - 24.1% using HPLC-MS/MRMand in the range of 4.3% - 13.1% with HPLC-MS/MRM3, for SP and NKA. The observedaccuracy was within 10% of the theoretical concentrations tested. HPLC-MS/MRM3 mayimprove the assay sensitivity to detect difference between samples by reducing significantly thepotential of interferences and therefore reduce instrumental errors.