Instrument-type effects on chemical isotope labeling LC-MS metabolome analysis: Quadrupole time-of-flight MS vs. Orbitrap MS

Abstract

Chemical isotope labeling (CIL) LC-MS is a powerful tool for metabolome analysis with markedly improved metabolomic coverage and quantification accuracy over the conventional LC-MS technique. In addition, with differential isotope labeling, each labeled metabolite is detected as a peak pair in the mass spectra, offering the possibility of differentiating true metabolite peaks from the singlet noise or background peaks. In this study, we examined the effects of instrument type on the detectability of true metabolites with a focus on the comparison of quadrupole time-of-flight (QTOF) and Orbitrap mass spectrometers. Using the same ultra-high-performance liquid chromatography setup and optimized running conditions for QTOF and Orbitrap, we compared the total number of peak pairs detected and identified from the two instruments using human urine and serum as the test samples. Many common peak pairs were detected from the two instruments; however, there were a significant number of unique peak pairs detected in each type of instrument. By combining the datasets obtained using QTOF and Orbitrap, the total number of peak pairs detected could be significantly increased. We also examined the effect of mass resolving power on peak pair detection in Orbitrap (60,000 vs. 120,000 resolution). The observed differences in peak pair detectability were much less than those of QTOF vs. Orbitrap. However, the type of peak pairs detected using different resolutions could be somewhat different, offering the possibility of increasing the overall number of peak pairs by combining the two datasets obtained at two different resolutions. The results from this study clearly indicate that instrument type can have a profound effect on metabolite detection in CIL LC-MS. Therefore, comparison of metabolome data generated using different instruments needs to be carefully done. Moreover, future research (e.g., hardware modifications) is warranted to minimize the differences in order to generate more reproducible metabolome data from different types of instruments.