Ultra high performance supercritical fluid chromatography coupled with mass spectrometry for simultaneous analysis of regio- and stereoisomers of triacylglycerols in natural fats and oils

Supercritical fluid chromatography (SFC) is a green analytical methods using non-toxic and inert CO₂ as the main mobile phase. The supercritical fluid has diffusion and viscosity comparable to that of gases, and the density is close to that of liquids. These physico-chemical properties lead to low back pressures allowing the use of high flow rates and good solubility properties when used as the mobile phase. Modern UHPSFC systems use columns with sub-2 mm particles as the stationary phase, providing higher separation efficiency, shorter analysis time, reduced energy consumption and use of harmful solvents. The dead volumes of the new SFC systems have been lowered while the reliability, quantitative performance and sensitivity have been improved.  Although (UHP)SFC has proven to be a powerful tool in both achiral and chiral lipid analyses, the potential of UHPSFC coupled with mass spectrometry (MS) in the chiral separation of lipids remains to be fully explored.

The aim of the present study was to explore the UHPSFC-ESI-MS to separate regio- and stereoisomers of triacylglycerols (TG) in complex matrices as natural fats and oils. The sixteen oils were extracted from plant sources. After solid phase extraction, the positional isomers of selected molecular species of the TG fractions were analysed with UHPSFC-MS using Daicel CHIRALPAK® IG-U (2 x 3.0 mm × 100 mm,1.6 μm) columns. Sn-PPO/POP/OPP and sn-POO/OPO/OOP were separated in twelve plant oils, whereas sn-OOS/SOO and sn-LLP/LPL/PLL were resolved and quantified in five and six oils, respectively.  Sn-POP is the major isomer within this TG species in all the plant oils analysed. Enantiomeric ratio within other TG species varied among the oils from different species. This study demonstrates UHPSFC-ESI-MS to be a powerful green analytical method for separating TG enantiomers in natural matrices as a highly demanding task of analytical chemistry.