Quantitative lipopolysaccharide analysis using HPLC/MS/MS and its combination with the limulus amebocyte lysate assay

Jean-Paul Pais de Barros (1, 2, 3) , Thomas Gautier (1, 2, 3) , Wahib Sali (1, 2, 3) , Christophe Adrie (4) , Hélène Choubley (3) , Emilie Charron (3) , Caroline Lalande (3) , Naïg Le Guern (1, 2, 3) , Valérie Deckert (1, 2, 3) , Mehran Monchi (5) , Jean-Pierre Quenot (1, 2, 3) , Laurent Lagrost (1, 2, 3, 6)

1 LNC - Lipides - Nutrition - Cancer (U866)
2 UBFC - Université Bourgogne Franche-Comté [COMUE]
3 LabEx LipSTIC - Laboratoire d'Excellence : Lipoprotéines et Santé : prévention et Traitement des maladies Inflammatoires et du Cancer
4 Service de physiologie et explorations fonctionelles [CHU Cochin]
5 Service de Médecine intensive [CH de Melun]
6 Service de recherche clinique [CHU de Dijon]

Quantitation of plasma lipopolysaccharides (LPSs) might be used to document Gram-negative bacterial infection. In the present work, LPS-derived 3-hydroxymyristate was extracted from plasma samples with an organic solvent, separated by reversed phase HPLC, and quantitated by MS/MS. This mass assay was combined with the limulus amebocyte lysate (LAL) bioassay to monitor neutralization of LPS activity in biological samples. The described HPLC/MS/MS method is a reliable, practical, accurate, and sensitive tool to quantitate LPS. The combination of the LAL and HPLC/MS/MS analyses provided new evidence for the intrinsic capacity of plasma lipoproteins and phospholipid transfer protein to neutralize the activity of LPS. In a subset of patients with systemic inflammatory response syndrome, with documented infection but with a negative plasma LAL test, significant amounts of LPS were measured by the HPLC/MS/MS method. Patients with the highest plasma LPS concentration were more severely ill. HPLC/MS/MS is a relevant method to quantitate endotoxin in a sample, to assess the efficacy of LPS neutralization, and to evaluate the proinflammatory potential of LPS in vivo.

Mots clés: diagnostic tool; human; inflammation; lipid transfer protein; lipoprotein; liquid chromatography tandem mass spectrometry; mass spectrometry; mouse; sepsis; systemic inflammatory response syndrome

Domaines: Biochimie, Biologie Moléculaire

Jean-Paul Pais de Barros, Thomas Gautier, Wahib Sali, Christophe Adrie, Hélène Choubley, et al.. Quantitative lipopolysaccharide analysis using HPLC/MS/MS and its combination with the limulus amebocyte lysate assay. Journal of Lipid Research, 2015, 56 (7), pp.1363 - 1369. ⟨10.1194/jlr.D059725⟩. ⟨hal-01687093⟩