Sphingosine-1-phosphate (S1P, HMDB00277) is a biologically significant lysophospholipid (specifically a sphingolipid), with important roles in intracellular inflammation, cancer, and Alzheimer’s disease1. Additionally, S1P is involved in calcium signaling pathway, cyclooxygenase-2 (COX-2) inductions, and eicosanoid production2. We have developed a quantitative LC-MS/MS assay, the S1P Panel, that quantifies four metabolites in sphingolipid metabolism (KEGG HSA00600): sphingosine-1-phosphate (green trace), sphingosine (pink trace), sphinganine (blue trace), and d18:2-S1P (yellow). A chromatogram of the analysis of a tissue sample is shown below; the total analysis time is five minutes per sample and carryover between samples was validated at <0.1%. While the utility and biological function of S1P, sphingosine, and sphinganine are well documented, the assay also quantifies d18:2-S1P, which has been shown as a reliable marker to identify samples which were improperly collected or stored at room temperature for extended periods of time3.
The panel has been validated for quantifying these compounds from variety of matrices including plasma, serum, and various tissues. The assay only requires 10 uL of serum or plasma and can be performed on <50 mg of tissue. The quantification range used is 0.2 uM to 125 uM. Analysis is performed using LC-MS/MS operating in multiple reaction monitoring mode and Skyline is used to quantify the metabolites based on external calibration curves (standards from Avanti Polar Lipids) with the use of d7-S1P as an internal standard. An example of a calibration curve shown below for sphingosine-1-phosphate. Please inquire to Dr. Will Thompson or Sarah Rains if you have an interest in this assay.
- Maceyka M, Harikumar KB, Milstien S, Spiegel S. SPHINGOSINE-1-PHOSPHATE SIGNALING AND ITS ROLE IN DISEASE. Trends in Cell Biology. 2012;22(1):50-60. doi:10.1016/j.tcb.2011.09.003.
- National Center for Biotechnology Information. PubChem Compound Database; CID=5283560, https://pubchem.ncbi.nlm.nih.gov/compound/5283560
- Liu X, Hoene M, Yin P, Fritsche L, Plomgaard P, Hansen J, Nakas C, Niess A, Hudemann J, Haap M, Mendy M, Weigert C, Wang X, Fritsche A, Peter A, Häring, H-U, Xu, G, and Lehmann R. QUALITY CONTROL OF SERUM AND PLASMA BY QUANTIFICATION OF (4E,14Z)-SPHINGADIENINE-C18-1-PHOSPHATE UNCOVERS COMMON PREANALYTICAL ERRORS DURING HANDLING OF WHOLE BLOOD. Clinical Chemistry. 2018;64:5. Doi:10.1373/clinchem.2017.277905.