Analysis of Phospholipids, Lysophospholipids, and Their Linked Fatty Acyl Chains in Yellow Lupin Seeds (Lupinus luteus L.) by Liquid Chromatography and Tandem Mass Spectrometry

Abstract

Hydrophilic interaction liquid chromatography (HILIC) and electrospray ionization (ESI) coupled to either Fourier-transform (FT) orbital-trap or linear ion-trap tandem mass spectrometry (LIT-MS/MS) was used to characterize the phospholipidome of yellow lupin (Lupinus luteus) seeds. Phosphatidylcholines (PC) were the most abundant species (41 ± 6%), which were followed by lyso-forms LPC (30 ± 11%), phosphatidylethanolamines (PE, 13 ± 4%), phosphatidylglycerols (PG, 5.1 ± 1.7%), phosphatidic acids (PA, 4.9 ± 1.8%), phosphatidylinositols (PI, 4.7 ± 1.1%), and LPE (1.2 ± 0.5%). The occurrence of both isomeric forms of several LPC and LPE was inferred by a well-defined fragmentation pattern observed in negative ion mode. An unprecedented characterization of more than 200 polar lipids including 52 PC, 42 PE, 42 PA, 35 PG, 16 LPC, 13 LPE, and 10 PI, is reported. The most abundant fatty acids (FA) as esterified acyl chains in PL were 18:1 (oleic), 18:2 (linoleic), 16:0 (palmitic), and 18:3 (linolenic) with relatively high contents of long fatty acyl chains such as 22:0 (behenic), 24:0 (lignoceric), 20:1 (gondoic), and 22:1 (erucic). Their occurrence was confirmed by reversed-phase (RP) LC-ESI-FTMS analysis of a chemically hydrolyzed sample extract in acid conditions at 100 °C for 45 min.

Identification and quantification of phospholipids in strawberry seeds and pulp (Fragaria × ananassa cv San Andreas) by liquid chromatography with electrospray ionization and tandem mass spectrometry

“…The lipidic extracts of both strawberry pulp and seeds were investigated in terms of quantitative content and regiochemistry of PL. This last aspect was accomplished by CID‐MS/MS of deprotonated molecules, [M − H] − , along with fragmentation patterns interpretation by either referring to standard compounds 17 or comparison with literature data 33–37 . As an example, Figure 2A reports the CID‐MS/MS spectrum obtained from a precursor ion [M − H] − at m/z 743.5.…”

“…Figure 4 displays the CID‐MS/MS spectrum of representative LPE 18:3 at m/z 474.3 (Figure 4A,B) and LPC 18:2 at m/z 504.3 (Figure 4C,D). HILIC silica‐based fused‐core column was employed to separate both lyso‐regioisomers, being sn ‐1 isomers more retained than sn ‐2 ones 17,31 . The XIC chromatograms at m/z 474.3 and m/z 504.3, with two isobaric peaks labelled as 1 and 2 corresponding to both isomers, are reported in the inset of Figure 4A,C, respectively.…”

“…A volume of 5 μL of strawberry seed or pulp lipid extract was injected into the column using the Autosampler (Thermo Scientific). An optimized binary elution program, 17 based on water and 2.5‐mmol L −1 ammonium formate (solvent A) and acetonitrile (solvent B), both containing 0.1% (v/v) of formic acid, was adopted: 0–5 min, linear gradient from 97% to 88% solvent B; 5–10 min, isocratic at 88% solvent B; 10–11 min, linear gradient from 88% to 81% solvent B; 11–20 min, linear gradient from 81% to 70% solvent B; 20–22 min, linear gradient from 70% to 50% solvent B; 22–28 min isocratic at 50% solvent B; 28–30 min, return to the initial composition, followed by a 5‐min equilibration time.…”

“…They also serve as a primary contact site between intracellular and extracellular spaces and between subcellular compartments. 32 17 or comparison with literature data. [33][34][35][36][37] As an example,…”

“…(A), PA 18:2/18:3 at m/z 693.5 and (B), isobaric PI 18:2/18:2 and 18:1/18.3 at m/z 857.5 18:3 at m/z 474.3 (Figure 4A,B) and LPC 18:2 at m/z 504.3 (Figure 4C, D). HILIC silica-based fused-core column was employed to separate both lyso-regioisomers, being sn-1 isomers more retained than sn-2 ones 17,31. The XIC chromatograms at m/z 474 3.…”