Liquid Chromatography–Light Scattering Detector–Mass Spectrometric Analysis of Digested Oxidized Rapeseed Oil

Abstract

Rapeseed oil was oxidized chemically and thermally to produce two distinct oxidized oils. These oils, along with unoxidized oils, were subjected to an artificial digestion model to simulate the digestive processes in humans. Lipid digestion involves lipases that break down the intact triacylglycerol (TAG) molecules first to diacylglycerols, and eventually to sn-2-monoacylglycerols (MAG) and free fatty acids. A high performance liquid chromatography-evaporative light scattering detector-electrospray ionization-mass spectrometric (HPLC-ELSD-ESI-MS) method was developed to monitor the lipolysis and the presence of oxidized lipids. The HPLC-ELSD-ESI-MS analysis enabled the separation and detection of nearly all the lipid species present in the sample after TAG hydrolysis. The HPLC-MS analyses of digestion products revealed that oxidized triacylglycerols are hydrolyzed by the digestive enzymes in a manner similar to that of native, unoxidized molecules. Significant amounts of sn-1(3)-MAG were found in all the samples after lipolysis, however, more of these were found in unoxidized rapeseed oil samples than in the oxidized oils. Several oxidized molecules were identified with the aid of synthesized oxylipids. This novel method is scalable to small-scale preparative fractionation of oxidized lipid molecules from a complex digestion sample. Also, the fingerprint-like, diagnostic, MS profiles of oxidized oils, reference compounds, and digestion products may be a great aid in comprehensive analysis of lipid oxidation and lipolysis.

A Simple Quantitative Approach for the Determination of Long and Medium Chain Lipids in Bio-relevant Matrices by High Performance Liquid Chromatography with Refractive Index Detection

“…Evaporative light-scattering detection (ELSD) for HPLC has been shown to be suitable for the separation of complex TAG mixtures and quantitative determination of mono-, di-and triacylglycerides (26,27). However, ELSD requires the use of very volatile solvents which may not be appropriate due to the potential volatility of low molecular weight lipids (28)(29)(30). Likewise, gas chromatography (GC) has been used to separate diacylglycerol and triacylglycerol molecular species (19).…”

Ultra high performance liquid chromatography–mass spectrometric analysis of oxidized free fatty acids and acylglycerols

“…Rapeseed oil was oxidized chemically and by heating in a convection oven (1008C, 48 h) to produce two distinct oxidized oils as described earlier [17]. Peroxide values of the chemically oxidized and oven oxidized oils were 20 and 100 mEq O 2 kg…”

“…This model has been described in detail elsewhere [17,21,22]. Briefly, a lipid free standard meal (900 mg) was mixed with the oil of interest (35 mg) and an internal standard (5 mg of 12-hydroxyoctadeanoic acid) in a test tube.…”

“…Temperature affects viscosity of the solvents and may be used as a tool to control the high back pressures. The gradient LC method was based on our earlier experiences [17] but numerous combinations of solvent composition, solvent gradients, flow rates, and temperatures were tested when developing the method. Table 1 shows comparison of chromatographic performance of the old HPLC method, the optimized UHPLC method and two other UHPLC methods tested during optimization.…”

“…Purpose of the current study was to improve significantly the speed of our earlier HPLC-ESI-MS method [17] for simultaneous analysis of free fatty acids (FFAs), monoacylglycerols (MAGs), diacylglycerols (DAGs), triacylglycerols (TAGs) and their oxidized equivalents. Effects of the elevated column temperature on resolution and back pressure were also studied.…”