Overview of Quantitative Lipid Analysis by Mass Spectrometry
www.lipidmaps.org
as performed by LIPID MAPS consortium on bone marrow derived macrophages (BMDM)
LIPID MAPS funded by Glue
Grant from :www.nigms.nih.gov
Extract bone marrow cells
Transfer to plates
Perform timecourse experiment
Repeat 3x
(replicates)
on plated cells
Aliquot samples for shipping to
core research labs
Fatty acids
Methanolic
HCl/Isooctane
extraction
Eicosanoids
Separate media from
cells
Glycerophospholipids
Methanolic HCl/CHCl3
extraction
Cardiolipins
Glycerolipids
Methanol/CHCl3
extraction
EtOAc/isooctane
extraction
SPE extraction of
media
Cholesteryl esters
EtOAc/isooctane
extraction
DFPI derivatization
of DAGS
Sphingolipids
Prenols
Saponification
Methanol/CHCl3 +
methanolic KOH
extraction
Methanol/CHCl3
extraction
Combination of LC-C18, LC-Si
and LC-NH2 separation
LC/MS analysis
Methanol/CHCl3
extraction
SPE extraction
GC/MS analysis
LC/MS analysis
LC/MS analysis
LC/MS analysis
LC/MS analysis
LC/MS analysis
Deuterated standards
(reverse phase)
(normal phase)
(normal phase)
(normal phase)
(normal phase)
ESI-QTRAP viaMRM methods
ESI-QTRAP
ESI-QTRAP
ESI-QTRAP
Deuterated standards
2-stage quantitation
ESI-QSTAR-XL using MS/MS
methods
Odd-chain standards
Odd-chain standards
[M+NH4]+ detection mode
[M+NH4]+/neutral loss
detection mode
Deuterated standards
Sterols
Deuterated standards
Combination of GC/MS,
LC/MS (reverse phase) on
ESI-QTRAP and APCI-MS
analysis
Deuterated standards
ESI-QTRAP and API-3000
Triple Quad detection with
MRM methods
(reverse phase)
QSTAR-XL via MRM methods
Nor-dolichol/CoQ6 standards
C12 analog standards
BIOINFORMATICSa,b
Data consolidation, normalization, statistical
analysis and databasing
Presentation in tabular and graphical formats
For details of extraction, purification and quantitation by MS, see:
Lipidomics reveals a remarkable diversity of lipids in human plasma.
Quehenberger O et al.,J Lipid Res 51, 3299-3305 (2010).
A mouse macrophage lipidome. Dennis EA et al., J Biol Chem 285, 3997639985 (2010)
Methods Enzymol. (Brown AH, ed.) 2007; Vol. 432 (multiple chapters)
LIPID MAPS Bioinformatics Corea, UCSD, 9500 Gilman Dr, La Jolla, CA 92093; Department of Bioengineeringb, UCSD, 9500 Gilman Dr, La Jolla, CA, 92093