title: Preanalytical Investigation of Polyunsaturated Fatty Acids and Eicosanoids in Human Plasma by Liquid Chromatography-Tandem Mass Spectrometry.
abstract: BACKGROUND
  Preanalytical variables have a great impact on sample matrices and are a source of laboratory errors. The effect of cryobanking, which is gaining great importance recently, requires systematic investigation. The arachidonic acid metabolism is useful as a quality marker since eicosanoids are easily subjected to in vitro oxidation processes.
  
  
  METHODS
  Polyunsaturated fatty acids (PUFAs) and related metabolites were analyzed by online solid-phase extraction coupled to liquid chromatography-tandem mass spectrometry. The influence of different plasma anticoagulants, as well as serum, freeze-thaw cycles (n = 5), short-term storage at 4°C, room temperature up to 120 minutes, and long-term storage at -20°C, -80°C, and -150°C up to 180 days, were investigated. We further investigated the influence of protein depletion, antioxidants, and shock-freezing on plasma.
  
  
  RESULTS
  PUFA metabolites were stable at 4°C in ethylenediaminetetraacetic acid (EDTA)-stabilized whole blood for 120 minutes and in EDTA-plasma for 30 minutes. Plasma stability at 4°C could be further increased up to 7 days after protein depletion, while addition of antioxidants such as butylated hydroxytoluene or coverage with nitrogen had no protective effects. Repeated freeze-thaw cycles (n > 1) resulted in eicosanoid formation up to 63%. Long-term storage at -20°C led to substantial eicosanoid increases after 30 days, which could be prevented by depleting proteins before storage. Cryobanking at -80°C and -150°C revealed decreased concentrations of eight eicosanoids after 180 days. An advantage of shock-freezing with liquid nitrogen could not be confirmed compared to conventional freezing.
  
  
  CONCLUSIONS
  Defined preanalytical conditions for eicosanoid analysis in human matrices are required to minimize in vitro data variability.
authors.affiliation: 1 Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University Hospital Leipzig , Leipzig, Germany .|2 LIFE-Leipzig Research Center for Civilization Diseases, Universität Leipzig , Leipzig, Germany .
authors.forename: Juliane
authors.initials: J
authors.lastname: Dorow
authors.affiliation: 1 Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University Hospital Leipzig , Leipzig, Germany .|2 LIFE-Leipzig Research Center for Civilization Diseases, Universität Leipzig , Leipzig, Germany .
authors.forename: Susen
authors.initials: S
authors.lastname: Becker
authors.affiliation: 1 Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University Hospital Leipzig , Leipzig, Germany .|2 LIFE-Leipzig Research Center for Civilization Diseases, Universität Leipzig , Leipzig, Germany .
authors.forename: Linda
authors.initials: L
authors.lastname: Kortz
authors.affiliation: 1 Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University Hospital Leipzig , Leipzig, Germany .|2 LIFE-Leipzig Research Center for Civilization Diseases, Universität Leipzig , Leipzig, Germany .
authors.forename: Joachim
authors.initials: J
authors.lastname: Thiery
authors.affiliation: 3 Department of Immunobiology, Institute of Biology, Universität Leipzig , Leipzig, Germany .
authors.forename: Sunna
authors.initials: S
authors.lastname: Hauschildt
authors.affiliation: 1 Institute of Laboratory Medicine, Clinical Chemistry and Molecular Diagnostics, University Hospital Leipzig , Leipzig, Germany .|2 LIFE-Leipzig Research Center for Civilization Diseases, Universität Leipzig , Leipzig, Germany .
authors.forename: Uta
authors.initials: U
authors.lastname: Ceglarek
mesh_terms: D000328:Adult; D000925:Anticoagulants; D002853:Chromatography, Liquid / Q000379:methods*; D004492:Edetic Acid; D015777:Eicosanoids / Q000097:blood*; D005231:Fatty Acids, Unsaturated / Q000097:blood*; D005260:Female; D006801:Humans; D008297:Male; D052616:Solid Phase Extraction; D053719:Tandem Mass Spectrometry / Q000379:methods*; D055815:Young Adult
chemical_list: D000925:Anticoagulants; D015777:Eicosanoids; D005231:Fatty Acids, Unsaturated; D004492:Edetic Acid