LC-MS/MS versus TLC plus GC methods: Consistency of glycerolipid and fatty acid profiles in microalgae and higher plant cells and effect of a nitrogen starvation
2017
Jouhet, Juliette | Lupette, Josselin | Clerc, Olivier | Magneschi, Leonardo | Bedhomme, Mariette | Collin, Séverine | Roy, Sylvaine | Maréchal, Eric | Rebeille, Fabrice | Physiologie cellulaire et végétale (LPCV) ; Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG) ; Direction de Recherche Fondamentale (CEA) (DRF (CEA)) ; Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)) ; Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA) | Commissariat à l'énergie atomique et aux énergies alternatives (CEA) | Institut National de la Recherche Agronomique (INRA) | Université Grenoble Alpes [2016-2019] (UGA [2016-2019]) | Centre National de la Recherche Scientifique (CNRS) | TOTAL-CEA ; CEA High Commissioner (Flagship program)
Methods to analyze lipidomes have considerably evolved, more and more based on massspectrometry technics (LC-MS/MS). However, accurate quantifications using these methodsrequire 13C-labeled standards for each lipid, which is not feasible because of the verylarge number of molecules. Thus, quantifications rely on standard molecules representativeof a whole class of lipids, which might lead to false estimations of some molecular species.Here, we determined and compared glycerolipid distributions from three different types ofcells, two microalgae (Phaeodactylum tricornutum, Nannochloropsis gaditana) and onehigher plant (Arabidopsis thaliana), using either LC-MS/MS or Thin Layer Chromatographycoupled with Gas Chromatography (TLC-GC), this last approach relying on the precisequantification of the fatty acids present in each glycerolipid class. Our results showed thatthe glycerolipid distribution was significantly different depending on the method used. Howcan one reconcile these two analytical methods? Here we propose that the possible biaswith MS data can be circumvented by systematically running in tandem with the sample tobe analyzed a lipid extract from a qualified control (QC) of each type of cells, previously analyzedby TLC-GC, and used as an external standard to quantify the MS results. As a casestudy, we applied this method to compare the impact of a nitrogen deficiency on the threetypes of cells.
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