Annotating unknown components from GC/EI-MS-based metabolite profiling experiments using GC/APCI(+)-QTOFMS

GC/EI-MS-based metabolite profiling of derivatized polar fractions of crude plant extracts typically reveals several hundred components. Thereof, only up to one half can be identified using mass spectral and retention index libraries, the rest remains unknown. In the present work, the utility of GC/APCI(+)-QTOFMS for the annotation of unknown components was explored. Hence, EI and APCI(+) mass spectra of ~100 known components were extracted from GC/EI-QMS and GC/APCI(+)-QTOFMS profiles obtained from a methoximated and trimethylsilylated root extract of Arabidopsis thaliana. Based on this reference set, adduct and fragment ion formation under APCI(+) conditions was examined and the calculation of elemental compositions evaluated. During these studies, most of the components formed dominating protonated molecular ions. Despite the high mass accuracy (|Δm| ≤ 3 mDa) and isotopic pattern accuracy (mSigma ≤ 30) the determination of a component’s unique native elemental composition requires additional information, namely the number of trimethylsilyl and methoxime moieties as well as the analysis of corresponding collision-induced dissociation (CID) mass spectra. After all, the reference set was used to develop a strategy for the pairwise assignment of EI and APCI(+) mass spectra. Proceeding from these findings, the annotation of unidentified components detected by GC/EI-QMS using GC/APCI(+)-QTOFMS and corresponding deuterated derivatization reagents was attempted. For a total of 25 unknown components, pairs of EI and APCI(+) mass spectra were compiled and elemental compositions determined. Integrative interpretation of EI and CID mass spectra resulted in 14 structural hypotheses, of which seven were confirmed using authenticated standards.