Chromatographic separation
of triacylglycerol (TG) enantiomers
is a highly challenging task of analytical chemistry because of the
similar physicochemical properties. The analysis of chiral TGs is
crucial for improving the knowledge of lipid biochemistry and for
understanding the nutritional properties of fats and oils. Thus, this
study aimed to systematically investigate the chiral resolution of
TGs consisting of three different fatty acyls (FAs). Thirty-three
asymmetric TG enantiopairs, including 49 synthesized enantiopure TGs
and racemic TGs, were analyzed with a recycling chiral HPLC system.
Twenty-six enantiopairs were successfully separated. Overall, having
both unsaturated and saturated FAs in the outer positions or a difference
in carbon chain length between two saturated FAs at the outer positions
favored the separation of enantiomers. The retention time at separation
correlated negatively with the sn-3 carbon number
of the early eluting enantiomer and positively with the carbon number
difference between sn-1 and sn-3.
When the samples were studied in separate groups based on unsaturation
and regioisomers, both the acyl carbon number and the degree of unsaturation
of FAs in all three positions influenced the separation and elution
behavior of chiral TGs, indicating an active role of both intermolecular
interactions and steric hindrances. This is the first systematic study
of the chiral separation of TGs consisting of three different FAs
using a large number of enantiopairs. The novel findings on the behavior
of TG enantiomers in a chiral environment provide important guidance
and reference for a stereospecific study of the chemistry and biochemistry
of natural lipids.