Robust Method for High-Throughput Screening of Fatty Acids by Multisegment Injection-Nonaqueous Capillary Electrophoresis–Mass Spectrometry with Stringent Quality Control
- Additional Document Info
- View All
High-throughput screening methods for fatty acid (FA) determination are urgently needed due to their critical biochemical roles in human health while serving as biomarkers of habitual diet and chronic disease risk assessment. Herein, we introduce multisegment injection-nonaqueous-capillary electrophoresis-mass spectrometry (MSI-NACE-MS) as a multiplexed separation platform for analysis of more than 20 nonesterified FAs in human serum or plasma. Optimization of experimental conditions was required to overcome major technical hurdles in MSI-NACE-MS prior to a rigorous method validation and intermethod comparison with gas chromatography/mass spectrometry (GC/MS). Following a simple methyl- tert-butyl ether extraction, seven serum extracts were analyzed directly by MSI-NACE-MS within a single run (<4 min/sample) under negative ion mode detection that incorporates stringent measures for quality control, including batch correction adjustment. Overall, excellent technical variance (RSD = 10%) and good mutual agreement was demonstrated for 12 nonesterified FAs consistently measured in 50 serum samples analyzed independently by MSI-NACE-MS and GC/MS within the same laboratory (mean bias = 24%, n = 600). Also, total hydrolyzed plasma FAs using MSI-NACE-MS was compared to mean concentrations reported from a NIST standard reference material as an interlaboratory method validation (mean bias = 15%, n = 20). Accurate prediction of ion migration behavior in CE also supports structural elucidation of FAs in conjunction with high resolution MS. For the first time, we demonstrate that MSI-NACE-MS offers a rapid yet robust platform for direct quantification of circulating FAs using volume-restricted blood specimens that expands metabolome coverage to encompass anionic classes of lipids as required for large-scale epidemiological studies.
has subject area