Unravelling ionization and fragmentation pathways of carotenoids using orbitrap technology: a first step towards identification of unknowns

Abstract

Vegetables are a major source of carotenoids and carotenoids are identified as potentially important natural antioxidants that may aid in the prevention of several human chronic degenerative diseases. Characterization of carotenoids in organic biological matrices is a crucial step in any research valorization trajectory. This study reports for the first time the use of high mass resolution and exact mass orbitrap technology for the elucidation of carotenoid fragmentation pathways. This contributes to the generation of new tools for identifying unknown carotenoids based on fragmentation patterns. Two different chromatographic methods making use of different mobile phases resulted in the generation of different ion species because of the large influence of the mobile phase solvent composition on ionization. It was shown that depending on the molecular ion species that are generated (protonated ions or radical molecular ions), different fragments are formed when applying higher energy collisional dissociation. Fragmentation and the abundance of fragments provide valuable structural information on the type of functional groups, the polyene backbone and the location of double bonds in ring structures of carotenoids. Furthermore, coherence between specific substructures in the molecules and characteristic fragmentation patterns was observed allowing the assignment of fragmentation patterns for carotenoid substructures that can theoretically be extrapolated to carotenoids with similar (sub)structures. Differentiation between isomeric carotenoids by compound specific fragments could however not be made for all the isomeric groups under study. As a wide variety of isomeric forms of carotenoids exist in nature, the combination of good chromatographic separation with high resolution mass spectrometry and other complementary qualitative structure elucidation techniques such as a photo diode array detector and/or nuclear magnetic resonance spectroscopy are indispensable for unambiguous identification of unknown carotenoids.