Performance Evaluation of Gas Chromatography–Atmospheric Pressure Chemical Ionization–Time-of-Flight Mass Spectrometry for Metabolic Fingerprinting and Profiling

Gas chromatography–atmospheric-pressure chemical ionization–time-of-flight mass spectrometry (GC–APCI–TOFMS) was compared to GC × GC–electron ionization (EI)–TOFMS, GC–EI–TOFMS, GC–chemical ionization (CI)–quadrupole mass spectrometry (qMS), and GC–EI–qMS in terms of reproducibility, dynamic range, limit of detection, and quantification using a mix of 43 metabolites and 12 stable isotope-labeled standards. Lower limits of quantification for GC–APCI–TOFMS ranged between 0.06 and 7.81 μM, and relative standard deviations for calibration replicates were between 0.4% and 8.7%. For all compounds and techniques, except in four cases, R 2 values were above 0.99. Regarding limits of quantification, GC–APCI–TOFMS was inferior to only GC × GC–EI–TOFMS, but outperformed all other techniques tested. GC–APCI–TOFMS was further applied to the metabolic fingerprinting of two Escherichia coli strains. Of 45 features that differed significantly (false discovery rate < 0.05) between the strains, 25 metabolites were identified through highly accurate and reproducible (Δm ± SD below 5 mDa over m/z 190–722) mass measurements. Starting from the quasimolecular ion, six additional metabolites were identified that had not been found in a previous study using GC × GC–EI–TOFMS and an EI mass spectral library for identification purposes. Silylation adducts formed in the APCI source assisted the identification of unknown compounds, as their formation is structure-dependent and is not observed for compounds lacking a carboxylic group.