Lipids are the macro biomolecules that are soluble in nonpolar solvents and make up the building blocks of the structure and function of living cells. Over the last two decades, lipids have come to be understood as far more than merely components of cellular membranes and forms of energy storage, and are now also being implicated to play important roles in a variety of diseases. Therefore, to gain insight into the metabolism and dynamics of lipids are crucial. Stable isotope-labeled lipids refer to one or more naturally occurring atoms in the molecule of the lipids are substituted with one or more stable isotopes of that atom, which are routinely used in lipidomic laboratories, especially in the analysis of lipid molecule metabolism and dynamics. They are considered ideal internal standards, and the gold standard for absolute quantitation.
Fig. 1 The common stable isotope-labeled lipids used in metabolic research.
The application of stable isotope-labeled lipids as tracer combined with mass spectrometric represents a perfect tool to study the lipid metabolism and dynamics of lipid species. Numerous studies have already been proved that. Some examples using stable isotope-labeled lipids to analyze the lipid metabolisms of fatty acids (FA), glycerophospholipids (GP) and sphingolipids (SL) are shown in below .
Metabolism of FA
The application of stable isotope-labeled FA as precursor is a common strategy to track the metabolism of longer chain FA (>16 carbons). For instance, the dietary supplementation of human volunteers with stable isotope-labeled FA is often used to follow and elucidate pathways of postprandial fat deposition or to determine FA flux in plasma. In addition, commercially available D3-labeled saturated FA are used in many studies to analyze the activity of desaturases. More studies of FA metabolism by using labeled FA are listed in Table 1.
Table 1. Studies of FA metabolism by using stable isotope-labeled FA.
|Experimental system and matrix
|[U–13C]-FA 16:0, TG-[U–13C]-16:0, D2-FA 16:0, TG-D2-FA 16:0
|[U–13C]-FA 16:0, D2-FA 16:0
|D3-FA 16:0, D3-FA 16:1, D3-FA 18:0, D3-FA
18:1n-9, D3-FA C18:1n-7
cells derived from human
|D5/D7- FA 16:0, D5/D7-FA 16:1, D5/D7-FA
18:0, D5/D7-FA 18:1
|D5-FA 18:0, D7-FA 18:0
Metabolism of GP
When the GP contain FA or their metabolites, stable isotope-labeled FA can be applied to monitor GP synthesis. For example, Tserng et al. studied the turnover of phosphatidylcholine and diacylglycerol in HL60 cells with [13C4]-palmitate . In addition, stable isotope-labeled GP are excellent tools to study GP remodeling according to Kainu et al. The authors introduced phospholipid species that were labeled on the head group into cultured cells via mediation by methyl-b-cyclodextrin and followed their remodeling by direct infusion ESI-MS/MS .
Metabolism of SL
labeled palmitate may be used to analyze the biosynthesis of SL. Palmitate may be incorporated both in the sphingoid base backbone and as N-linked FA. And the label positions can be determined by mass spectrometry. And then analysis of isotopic enrichment in the palmitoyl-CoA pool can calculate the rate of de novo SL synthesis. Similarly, labeled SL species may be applied to monitor SL transport and metabolism. Tserng et al.  used ceramide d18:1/[13C4]-16:0 to investigate the uptake and metabolism of this long chain ceramide in HL60 cells. More studies of SL metabolism by using labeled lipids are listed in Table 2.
Table 2. Studies of SL metabolism by using stable isotope-labeled lipids.
|Total Sph content
| Ceramide, HexCer, sphingomyelin
| [U–13C]-FA 16:0
|GC–MS of Cer
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- Ecker, J. and Liebisch, G. Application of stable isotopes to investigate the metabolism of fatty acids, glycerophospholipid and sphingolipid species[J]. Progress in Lipid Research, 2014, 54:14-31.
- Tserng, K.Y. and Griffin R.L. Phosphatidylcholine de novo synthesis and modification are carried out sequentially in HL60 cells: evidence from mass isotopomer distribution analysis[J]. Biochemistry 2004, 43:8125-35.
- Kainu V., et al. Electrospray ionization mass spectrometry and exogenous heavy isotope-labeled lipid species provide detailed information on aminophospholipid acyl chain remodeling[J]. J. Biol. Chem. 2008, 283:3676-87.
- Tserng K.Y. and Griffin R.L. Ceramide metabolite, not intact ceramide molecule, may be responsible for cellular toxicity[J]. Biochem. J. 2004, 380:715-22.
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