Direct and rapid profiling of triacylglycerols in extra virgin olive oil using UHPLC-MS/MS

Extra virgin olive oil (EVOO) prices reached record levels in 2023, due to the decline in global production, mainly linked to drought in the Mediterranean. Without effective controls, the already widespread fraud in this sector will irrevocably worsen. However, the reliability of the current standard method (CSM) for controlling the authenticity of EVOO has been questioned. Besides, authors have recently called for the replacement of certain steps to reduce analysis time and solvent consumption. In fact, CSM based on HPLC-RID analysis of triacylglycerols (TAGs) and GC analysis of fatty acid methyl esters (FAMEs) is tedious and very time consuming. Here, we optimized and validated a UHPLC-APCI-MS/MS method capable of unambiguously identifying 33 TAGs in EVOO in less than 6 min. The limit of detection calculated for the available TAG standards is between 0.3 and 2.5 ng/ml (0.02 ng-0.13 ng of injected quantity). The linearity of the method in real samples is satisfactory (r(2) > 0.99, n = 6) for 27 TAGs with an average area greater than 0.04 %. In real samples, the between-run precision (RSD %) for 29 TAGs with mean area greater than 0.02% does not exceed 11.6%, provided that the injected amount is equal to or greater than 4 mu g of EVOO sample. The application of this method to eleven EVOO samples, including a control sample from the International Olive Council, and the processing of the data with a PCA model allow their fine discrimination according to their TAG profiles. In addition to eliminating the GC step of FAME analysis, the high productivity of the proposed method significantly reduces the cost of this essential control step compared to CSM.