Identification of very long-chain (>C24) fatty acid methyl esters using gas chromatography coupled to quadrupole/time-of-flight mass spectrometry with atmospheric pressure chemical ionization source

Gas chromatography coupled to a quadrupole time-of-flight mass analyzer (QTOF) with an atmospheric pressure chemical ionization (APCI) source has been tested to study the ionization and mass spectrometric behavior of long-chain and very long-chain polyunsaturated fatty acids (LC-PUFAs, C18–24; VLC-PUFAs, >C24). The protonated molecule ([M+H]+), measured at accurate mass, became the base peak of the spectrum for all the studied compounds and was promoted by the addition of water into the source. This fact overcame the existing difficulties for the identification of VLC-PUFAs when using an electron ionization source (EI). The extensive fragmentation of PUFAs in this source is the main drawback due to the fact that since reference standards are not commercially available, final identification relies on retention time estimation. The application of GC−APCI-QTOF to the screening of lipid extracts from the eyes of different fish species added confidence to the identification of several VLC-PUFAs. Further investigation of ion ratios allowed to predict the position of key double bonds enabling the classification of VLC-PUFAs as ω3 or ω6 compounds. VLC-PUFAs spectra found in biological samples were compared to those obtained from corresponding peaks found in heterologous expression experiments of fish's Elovl4.The authors acknowledge the financial support of the Programa Estatal de Investigación, Desarrollo e Innovación orientada a los retos de la sociedad (Ministry of Economy and Competitiveness): Research Project “Ácidos grasos poliinsaturados de cadena muy larga en peces: biosíntesis e implicaciones durante fases tempranas de desarrollo en especies cultivadas”(AGL2013-40986-R). This work has been developed within the framework of the Research Unit of Marine Ecotoxicology (IATS-CSIC/IUPA-UJI). Tania Portolés acknowledges Juan de la Cierva Incorporation Program from Ministry of Economy and Competitiveness (IJCI-2014-20588) for funding her research.Peer reviewe