title: Hyphenating size-exclusion chromatography with electrospray mass spectrometry; using on-line liquid-liquid extraction to study the lipid composition of lipoprotein particles.
abstract: BACKGROUND
  Lipoproteins belong to the most commonly measured clinical biochemical parameters. Lipidomics is an orthogonal approach and aims to profile the individual lipid molecules that jointly form the lipoprotein particles. However, in the first step of the extraction of lipid molecules from serum, an organic solvent is used leading to dissociation of the lipoproteins. Thus far it has been impossible to combine lipidomics and lipoprotein analysis in one analytical system.
  
  
  METHODS
  Human plasma was diluted in phosphate-buffered saline (PBS) and injected onto a Superose 6 PC 3.2 column with PBS as a mobile phase to separate lipoproteins. The eluent was led to a Syrris FLLEX module, which also received CHCl3 /MeOH (3:1). The two phases were mixed and subsequently separated using a Teflon membrane in an especially designed pressurized flow chamber. The organic phase was led to a standard electrospray source of an Orbitrap mass spectrometer.
  
  
  RESULTS
  Size-exclusion chromatography (SEC) has been commonly applied to separate lipoproteins and is considered a practical alternative to ultracentrifugation. Through the on-line liquid-liquid extraction method it becomes possible to obtained detailed mass spectra of lipids across different lipoprotein fractions. The extracted ion chromatograms of specific lipid signals showed their distribution against the size of lipoprotein particles.
  
  
  CONCLUSIONS
  The application of on-line liquid-liquid extraction allows for the continuous electrospray-based mass spectral analysis of SEC eluent, providing the detailed lipid composition of lipoprotein particles separated by size. This approach provides new possibilities for the study of the biochemistry of lipoproteins.
authors.affiliation: Medical Research Council Human Nutrition Research, Elsie Widdowson Laboratory, 120 Fulbourn Road, Cambridge, CB1 9NL, UK.
authors.forename: Michael
authors.initials: M
authors.lastname: Osei
authors.affiliation: Medical Research Council Human Nutrition Research, Elsie Widdowson Laboratory, 120 Fulbourn Road, Cambridge, CB1 9NL, UK.
authors.forename: Julian L
authors.initials: JL
authors.lastname: Griffin
authors.affiliation: Medical Research Council Human Nutrition Research, Elsie Widdowson Laboratory, 120 Fulbourn Road, Cambridge, CB1 9NL, UK.
authors.forename: Albert
authors.initials: A
authors.lastname: Koulman
mesh_terms: D001331:Automation; D002850:Chromatography, Gel / Q000379:methods*; D006801:Humans; D008074:Lipoproteins / Q000737:chemistry* / Q000302:isolation & purification*; D059625:Liquid-Liquid Extraction / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D008074:Lipoproteins