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Development and validation of a LC-MS/MS method for determination of eight PEth homologues in whole blood

2022Dissertação de mestrado Acesso aberto
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datacite.subject.fosDepartamento de Biologia Animalpt_PT
dc.contributor.advisorBerg, Thomas
dc.contributor.advisorRebelo, Maria Teresa Ferreira Ramos Nabais de Oliveira, 1964-
dc.contributor.authorMaria, Marisa Henriques
dc.date.accessioned2022-03-30T09:15:11Z
dc.date.available2022-12-30T01:31:13Z
dc.date.issued2022
dc.date.submitted2021
dc.descriptionTese de mestrado, Biologia Humana e Ambiente, Universidade de Lisboa, Faculdade de Ciências, 2022pt_PT
dc.description.abstractPhosphatidylethanol (PEth) is a group of direct ethanol biomarkers used to distinguish between different drinking patterns, such as heavy and social drinkers. So far, the PEth homologue most used for this purpose is PEth 16:0/18:1. This homologue is also the one determined in most of the LC-MS/MS methods developed for PEth analysis. Often one or a few PEth homologues are quantified in LC-MS/MS methods published, but recently a LC-MS/MS method for the determination of six PEth homologues was developed. The main aim of the dissertation was to develop a LC-MS/MS method for the quantitative determination of eight of the most common PEth homologues in whole blood. This can be helpful, especially for the determination of PEth in blood from people with high alcohol consumption. Chromatographic separation of the PEth homologues was tested on five UHPLC columns. Different mobile phases were investigated to optimize separation. Best chromatographic separation of the PEth homologues was achieved using an Acquity BEH C18 column (100 2.1 mm I.D., 1.7 µm particles) and a mobile phase consisting of 5 mM ammonium acetate pH 5 (Solvent A) and methanol (Solvent B). The total run time was 5.7 minutes. MS/MS parameters were optimized for each of the eight PEth homologues and internal standards. Two MRM transitions ions, those with the highest responses, were chosen for each compound. Isotope labeled internal standards were used for five of the eight PEth homologues. Two sample preparation techniques, liquid-liquid extraction (LLE) and 96-well supported liquid extraction (SLE), were investigated to extract the PEth homologues from blood. The extraction recoveries obtained were within 45-82% when LLE was used and within 68-80% when 96-well SLE was used. Matrix effects were studied and showed ion suppression for the majority of the PEth homologues, and ion enhancement for PEth 17:0/18:1. For compounds with their own internal standards, the matrix effects were corrected for their internal standard. An LC-MS/MS method for the determination of eight PEth homologues was developed and validated. The validation showed that UHPLC-MS/MS method is precise, accurate, and sensitive for its purposes.pt_PT
dc.identifier.tid202994511
dc.identifier.tid202994511pt_PT
dc.identifier.urihttp://hdl.handle.net/10451/52084
dc.language.isoengpt_PT
dc.subjectPEth homólogospt_PT
dc.subjectLC-MS/MSpt_PT
dc.subjectExtração líquido-líquido (LLE)pt_PT
dc.subjectExtração sólido-líquido em 96-well (96-well SLE)pt_PT
dc.subjectValidação do métodopt_PT
dc.subjectTeses de mestrado - 2022pt_PT
dc.titleDevelopment and validation of a LC-MS/MS method for determination of eight PEth homologues in whole bloodpt_PT
dc.typemaster thesis
dspace.entity.typePublication
rcaap.rightsopenAccesspt_PT
rcaap.typemasterThesispt_PT
thesis.degree.nameTese de mestrado em Biologia Humana e Ambientept_PT

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