Alternative mass spectrometry techniques for the validation of the fragmentation pattern of capsaicin and dihydrocapsaicin

Abstract

Rationale Capsaicinoids are prevalent secondary metabolites in many natural and synthetic pharmacological compounds. To date, several soft ionization studies related to capsaicinoids have been reported; they all proposed a common fragmentation pattern based on a rearrangement of the aromatic double bonds and the fragmentation of the various positional acyl chains. However, the mechanism has never been validated by high‐resolution analyses. Consequently, in this work, a validated fragmentation mechanism of the main capsaicinoids, capsaicin (1) and dihydrocapsaicin (2), is offered. Methods In order to propose and validate a common electron ionization (EI) fragmentation mechanism for the target analytes, the following mass spectrometric methods were employed: collision‐induced dissociation (CID) by means of linked scans (LS), reinforcing the methodology by high‐resolution mass spectrometry (HRMS), in addition to appropriate deuterium‐labeled experiments performed using gas chromatography/mass spectrometry (GC/MS) and direct analysis in real time (DART). Results In a first stage, a common EI fragmentation pattern comprising two pathways was proposed for compounds 1 and 2; then, the suggested mechanism was validated by CID‐LS together with HRMS complemented by DART‐deuterium‐labeling studies. The obtained results are indicative that the corresponding molecular ions were conveniently observed, m/z 305 and m/z 307; it is worth noting that the common base peak is in correspondence with a tropylium ion derivative (m/z 137), as a consequence of a McLafferty rearrangement. In addition to these highlighted fragments, other common ions, m/z 122 and m/z 94, and their corresponding trajectory, were confirmed using the same approach. Finally, the proposed mechanism was complementarily validated by deuterium‐labeling studies, taking into account the two exchangeable hydrogens present in the phenolic and the amidic moieties. Conclusions A common validated EI fragmentation pattern for both capsaicin and dihydrocapsaicin was established using appropriated mass spectrometric methods together with convenient hydrogen/deuterium labeling. This study provides a new alternative to validate mechanisms of fragmentation of important natural products.

Differentiation of 2‐hydroxyglutarate enantiomers and its lactones by gas chromatography/electron ionization tandem mass spectrometry

“…The same experiment was repeated for the 2‐hg lactone confirming that 117 m/z was not a major fragment of the 2‐hg lactone and that 85, 57, and 29 m/z were shared fragments with the acyclic compound (Figures C and D). Some studies advocate for the use of high‐resolution GC/MS, such as GCTOFMS/GCOrbitrap, to have unambiguous identification of fragments . Though imperative with larger and more complex molecules, the extra resolution would not provide any additional data given that the typical fragments observed and expected for 2‐hg methyl esters did not result in fragments close in m/z , nor would the analysis benefit from the determination of mass defect differences.…”

“…Some studies advocate for the use of high-resolution GC/MS, such as GCTOFMS/GCOrbitrap, to have unambiguous identification of fragments. [16][17][18] Though imperative with larger and more complex molecules, the extra resolution would not provide any additional data given that the typical fragments observed and expected for 2-hg methyl esters did not result in fragments close in m/z, nor would the analysis benefit from the determination of mass defect differences. The SRM analysis by QqQ provided the necessary specificity to differentiate the EI fragments of 2-hg.…”

Z0 ion from saikosaponins with 16 α‐OH and Y0‐H2O from saikosaponins with 16 β‐OH may underlie their different dissociation patterns of [aglycone − H2O + H]+

“…This may shed light on characterizing other epimeric saponins in future studies and facilitate the characterization and identification of the metabolites of epimeric saponins in both in vivo and in vitro experiments. However, the mechanism of the stereospecific fragmentations of epimers does need further study, in which the isotope‐labelled method and density functional theory or quantum chemistry calculations may play a key role.…”

Rapid, efficient, and accurate determination of aflatoxins and capsaicinoids in vegetable oils by immunomagnetic sorbents coupled with UHPLC–MS/MS

“…Those immunochromatographic techniques mainly focused on the total amount of AFTs and capsaicinoids, but few reported about the component and distribution of those analytes in vegetable oils. Chromatography instruments consist of ultra‐performance liquid/gas separation system coupled with ultraviolet, diode assay detector, tandem mass spectrometry (MS/MS), and high‐resolution MS (Fan et al., 2017; Ma et al., 2016; Rosas et al., 2019). Owing the simplicity, rapidity, and high resolution, ultra‐performance liquid chromatography coupled with tandem MS (UHPLC–MS/MS) is widely utilized as the in‐lab and official method for the accurate and sensitive detection of AFTs and capsaicinoids in vegetable oils.…”