Abstract
In the present study the running conditions (sample pH, scan number, relaxation delay and NMR solvent) for the quantification of phospholipids by 31P NMR using commercial pure standards were optimised. A mixture of pure standards was run at different pH (7.3, 7.8, 9.5 and 12), scan numbers (64, 128, 192 and 512), relaxation delay times (1s, 2s, 3.5s and 10s) and NMR solvents (D2O/EDTA/sodium cholate or CDCl3/CD3OD/EDTA) to achieve this goal. Best 31P NMR conditions were found to be pH 7.3, a scan number of 192 and a relaxation delay of 3.5s, which were used for the quantification of natural samples (Krill oil, soybean lecithin and egg phospholipid) in D2O/EDTA/sodium cholate or CDCl3/CD3OD/EDTA. Phospholipids and lysophospholipids were highly resolved and detected in D2O/EDTA/sodium cholate compared to CDCl3/CD3OD/EDTA. A number of phospholipids (lysophoshatidylglycerol-2, LPG-2; lysophosphatidylethanolamine, LPE; N-acyl phosphatidylethanolamine, APE) were not detected in CDCl3/CD3OD/EDTA, but they were detected in D2O/EDTA/sodium cholate. Therefore, it is concluded that D2O/EDTA/sodium cholate is a better detection solvent for 31P NMR quantification of phospholipids extracted from natural sources.
•31P NMR running conditions were optimised for phospholipid analysis.•Phospholipids were hydrolysed under alkaline conditions.•Soybean lecithin, krill oil and egg phospholipids were analysed in two NMR solvents.•D2O provided better detection and quantification of phospholipids than CDCl3.