Quantitation of triacylglycerols in plant oils using HPLC with APCI‐MS, evaporative light‐scattering, and UV detection

Abstract

The main constituents of plant oils are complex mixtures of TGs differing in acyl chain lengths, number and positions of double bonds, and regioisomerism. A non-aqueous reversed-phase HPLC method with acetonitrile-2-propanol gradient and 30 + 15 cm NovaPak C18 columns makes possible an unambiguous identification of the highest number of TGs ever reported for these oils, based on positive-ion APCI mass spectra. A new approach to TG quantitation is based on the use of response factors with three typical detection techniques for that purpose (APCI-MS, evaporative light-scattering detection, and UV at 205 nm). Response factors of 23 single-acid TGs (saturated TGs from C7 to C22, 7 unsaturated TGs), 4 mixed-acid TGs, diolein and monoolein are calculated from their calibration curves and related to OOO. Due to differences between saturated and unsaturated acyl chains, the use of response factors significantly improves the quantitation of TGs. 133 TGs containing 22 fatty acids with 8-25 carbon atoms and 0-3 double bonds are identified and quantified in 9 plant oils (walnut, hazelnut, cashew nut, almond, poppy seed, yellow melon, mango, fig, date) using HPLC/APCI-MS with a response factor approach. Average parameters and relative fatty acid concentrations are calculated with both HPLC/APCI-MS and GC/ FID.

Theory and Practice of UHPLC and UHPLC–MS

“…Temperature plays an important role in the optimization of the chromatographic separation of lipids, which is not limited to Ag-HPLC, but is valid for other separation modes as well, such as NARP (Holcapek et al, 1999(Holcapek et al, , 2005Lísa and Holcapek, 2008). Increased temperature can result in the loss of resolution for critical TG pairs in NARP mode, where the retention time depends inversely on the temperature.…”

“…Ag-HPLC has the ability to resolve TG regioisomers. At present, the following techniques are the most convenient for the regioisomeric analysis: (1) Ag-HPLC Adlof and List, 2004;Adlof, 1995Adlof, , 2007Dugo et al, 2004Lísa et al, 2009a,b;, (2) MS (Holcapek et al, , 2005Lísa and Holcapek, 2008;Leskinen et al, 2008;Byrdwell, , 2005Byrdwell et al, 1996;Lísa et al, 2009a,b;Mottram et al, 1997Mottram et al, , 2001Mottram and Evershed, 1996), (3) enzymatic reactions (e.g., pancreatic lipase, phospholipase A 2 ) followed by some analytical technique (e.g., Ag-HPLC), and (4) nuclear magnetic resonance (NMR) spectroscopy (Standal et al, 2009). Each technique has certain limitations.…”

“…Mass spectra of TG provide two basic types of ions sufficient for the structural assignment (Byrdwell et al, 1996;Mottram et al, 1997;: (1) molecular adducts with protons (2) Mottram et al, 1997Mottram et al, , 2001, electrospray ionization (Malone and Evans, 2004;Li and Evans, 2005;Li et al, 2006;Gakwaya et al, 2007;Herrera et al, 2010;Leveque et al, 2010), and matrix-assisted laser desorption/ionization (Pittenauer and Allmaier, 2009 Mottram et al, 1997Mottram et al, , 2001Holcapek et al, , 2005Cvacka et al, 2006;Lísa et al, 2007aLísa et al, ,b, 2009aLeskinen et al, 2008), as illustrated in the example of regioisomers SLnO/SOLn/OSLn in Fig. 4.8.…”

“…4.8. The simple approach just determines the prevailing FAs in the middle sn-2 position based on the lower relative abundance of the corresponding [M þ HeR i COOH] þ fragment ion Mottram et al, 1997Mottram et al, , 2001Dugo et al, 2004;Lísa and Holcapek, 2008), but the type of FAs (mainly DB number and positions) (Holcapek et al, 2005;Li and Evans, 2005;Li et al, 2006;Gakwaya et al, 2007) also affects relative abundances of corresponding [M þ HeR i COOH] þ fragment ions. An interesting example of the strong effect of DB position is the ratio of […”

Quantification of Triacylglycerol Molecular Species in Cocoa Butter Using High-Performance Liquid Chromatography Equipped with Nano Quantity Analyte Detector

“…An MS operated in selected-ion-recording or multiple-reaction-monitoring mode has a wide range of linearity that enables highly selective detection of specific TAG molecular species. However, these detection modes give different intensities for each type of TAG molecular species 22 . Although the corona charged aerosol detector CAD has been considered for the quantification of TAG molecular species, the obtained calibration curve is not linear 23 .…”