Identification and Nontargeted Screening of Phytosterol Esters in Oilseeds and Oils Based on Fragmentation Characteristics via Ultrahigh–Performance Liquid Chromatography High–Resolution Mass Spectrometry

Phytosterol (PS) esters are natural bioactive compounds with various physiological functions, such as reducing cholesterol levels and protecting cardiovascular health. In the present study, a novel method for identifying and screening phytosterol esters (PSEs) was developed by ultrahigh–performance liquid chromatography coupled with high–resolution mass spectrometry. First, the structural characteristics, mass spectrometry behaviors, and fragmentation rules of PSEs were investigated and summarized. An in–house theoretical database was then established. The database contained 1,176 PSEs, including their chemical formulas, exact molecular masses, protonated forms, accurate masses of precursor ions, and characteristic fragment ions. A nontargeted screening workflow was then developed to identify unknown PSEs and distinguish isomers with different fatty acid acyl chain lengths based on precursor ions and characteristic fragments. To validate the applicability of the method, the most effective extraction method (Matyash) for PSEs was determined, and this nontargeted screening was applied to 23 types of oilseeds and oils from China. Ultimately, 12 PSEs were identified, and their compositional characteristics in these samples were analyzed. This innovative approach not only advances the analytical techniques for PSEs in oilseeds and oils but also lays a foundation for future research on the nutritional profiling of various food substrates, potentially facilitating the development of functional foods aimed at enhancing human health.