Nonesterified Fatty Acid Determination for Functional Lipidomics: Comprehensive Ultrahigh Performance Liquid Chromatography–Tandem Mass Spectrometry Quantitation, Qualification, and Parameter Prediction
Autor: | Martina Weber, Wolfgang Peissner, Christian Hellmuth, Berthold Koletzko |
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Rok vydání: | 2012 |
Předmět: |
0303 health sciences
Chromatography Chemistry 010401 analytical chemistry Water Fatty Acids Nonesterified Tandem mass spectrometry Mass spectrometry 01 natural sciences High-performance liquid chromatography 0104 chemical sciences Analytical Chemistry Triple quadrupole mass spectrometer 03 medical and health sciences NEFA Tandem Mass Spectrometry Liquid chromatography–mass spectrometry Lipidomics Humans Sample preparation Chromatography High Pressure Liquid 030304 developmental biology |
Zdroj: | Analytical Chemistry. 84:1483-1490 |
ISSN: | 1520-6882 0003-2700 |
Popis: | Despite their central importance for lipid metabolism, straightforward quantitative methods for determination of nonesterified fatty acid (NEFA) species are still missing. The protocol presented here provides unbiased quantitation of plasma NEFA species by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Simple deproteination of plasma in organic solvent solution yields high accuracy, including both the unbound and initially protein-bound fractions, while avoiding interferences from hydrolysis of esterified fatty acids from other lipid classes. Sample preparation is fast and nonexpensive, hence well suited for automation and high-throughput applications. Separation of isotopologic NEFA is achieved using ultrahigh-performance liquid chromatography (UPLC) coupled to triple quadrupole LC-MS/MS detection. In combination with automated liquid handling, total assay time per sample is less than 15 min. The analytical spectrum extends beyond readily available NEFA standard compounds by a regression model predicting all the relevant analytical parameters (retention time, ion path settings, and response factor) of NEFA species based on chain length and number of double bonds. Detection of 50 NEFA species and accurate quantification of 36 NEFA species in human plasma is described, the highest numbers ever reported for a LC-MS application. Accuracy and precision are within widely accepted limits. The use of qualifier ions supports unequivocal analyte verification. |
Databáze: | OpenAIRE |
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