Method development for the separation of phospholipids by subcritical fluid chromatography

• Published in: Journal of Chromatography A Vol. 826; no. 2; pp. 241 - 247
• Main Authors: Eckard, Phyllis R, Taylor, Larry T, Slack, Gregory C
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 27.11.1998; Elsevier
• Subjects: Analytical chemistry; Analytical, structural and metabolic biochemistry; Biological and medical sciences; Chemistry; Chromatographic methods and physical methods associated with chromatography; Exact sciences and technology; Fundamental and applied biological sciences. Psychology; Glycerolipids, phospholipids; Lipids; Other biological molecules; Other chromatographic methods; Phospholipids; Phospholipids; Subcritical fluid chromatography; Analysis method; Light scattering; Phospholipid; Qualitative analysis; Detection; Complex lipid; Supercritical fluid chromatography
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(98)00723-7

A packed-column subcritical fluid chromatographic separation of five phospholipids was achieved in less than 20 min by using a Luna octyl column (25 cm×4.6 mm I.D., 5 μm) employing evaporative light scattering detection. The chromatographic conditions used for the separation of (a) 1,2-dipalmitoyl- sn-glycero-3-phosphate (DPPA), 1,2-dipalmitoyl- sn-glycero-3-phosphocholine (DPPC), (c) poly(ethylene glycolated)-1,2-dipalmitoyl- sn-glycero-3-phosphocholine (DPPE–PEG), (d) 1,2-dicaproyl- sn-glycero-3-phosphocholine (DCPC) and (e) 1,2-dipalmitoyl- sn-glycero-3-[phospho- rac-(1-glycerol)] (DPPG) were: 9% ethanol–methanol (50:50, v/v), containing 0.10% (v/v) trifluoroacetic acid for 2 min, increasing by 0.5%/min to 15% in 14 min, to 40% at 14.1 min, at a column temperature of 70°C, an outlet pressure of 135 bar and a mobile phase flow-rate of 2.0 ml/min. The separation was shown to be significantly influenced by the presence of a mobile phase acidic additive (e.g. trifluoroacetic acid) and its concentration, the overall modifier ramp rate and the column outlet pressure.