Polar lipids and other metabolites extracted from plasma from lean, diabetic, and obese Zucker rats were analyzed by flow
injection using high resolution time of flight mass spectrometry. Metabolites were confidently identified and differences
in the metabolite profiles determined.
The need for high-throughput analysis of biological samples is of growing importance owing to its clinical utility and the
ability to leverage new developments in technology. This is particularly true in metabolomics where screening to evaluate
significant differences in key metabolites and metabolite families can be selectively observed using new tools. One such tool
is high performance time of flight mass spectrometry. The mass accuracy and resolving power now offered allows for confident
evaluation of metabolites with little sample preparation.
Samples and Analysis
Rat plasma (Zucker; Bioreclamation) was precipitated with 80% cold methanol and the supernatant analyzed directly. Polar lipids
in the extracts were analyzed by injection into a stream of acetonitrile/water with formic acid.
Table I: Representative analytes detected in samples with mass error and change (Delta) in plasma from obese vs. lean rats
Mass Spectrometer – Citius™ LC-HRT (LECO Corporation, Saint Joseph, MI)
Mass Range – 240–940
Acquisition Conditions – 1 spectra/s; R =100,000; positive ion mode
Solvent delivery Conditions – 60 ÁL /min (acetonitrile/water/0.1% formic acid)
Injection Volume – 2 ÁL
Acquisition time – 2 min
The analyses provided a rich spectrum which can be background subtracted to yield useful information on the lipid content
of the simple plasma extract. This is shown in Figure 1 for data acquired at 100,000 resolving power. The resolving power
allows for separation of the M+H+2 isotope and M+H of unsaturated forms as shown for PCs containing 34 carbons. This requires
a resolving power in excess of 80,000.
Figure 1: Mass spectrum at R > 100,000 from plasma extract – (top) full acquisition; (bottom) M+2 isotope of PC34:2 and the
M of PC34:1 resolved.
The direct analysis of lipids using high performance time of flight provides a quick and efficient tool to screen differences
in lipids in complex extracts. The analysis time of 2 min provides for a rapid evaluation with the high resolving power allowing
for selective analysis.
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