US EPA

1040255 

Journal Article 

Simultaneous determination of sphingosine and sphingosine 1-phosphate in biological samples by liquid chromatography-tandem mass spectrometry 

Lan, T; Bi, H; Liu, W; Xie, X; Xu, S; Huang, H 

2011 

Yes 

Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
ISSN: 1570-0232
EISSN: 1873-376X 

879 

7-8 

520-526 

English 

21292568 

10.1016/j.jchromb.2011.01.015 

D-erythro-sphingosine (Sph) and its phosphorylated product, d-erythro-sphingosine 1-phosphate (S1P) are sphingolipids mediating numerous cellular processes. Imbalance of Sph/S1P levels contributes to many diseases. Given the interconversion of these two opposing signaling molecules, it is essential to examine their levels simultaneously. In the present study, we developed a rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to simultaneously quantify the levels of Sph and S1P in biological samples using C17-Sph and C17-S1P as internal standards. With one step of methanol-induced protein precipitation, each sample was subjected to LC-MS/MS analysis using positive electrospray ionization under selected reaction monitoring mode. The running time was within 4 min with a simple mobile phase consisting of methanol-0.1% formic acid (95:5, v/v) at a flow rate of 0.2 mL/min. Standard curves were linear over ranges of 1-100 ng/mL for Sph and 0.1-10 ng/mL for S1P with correlation coefficient (r²) greater than 0.997. The lower limit of quantifications (LLOQs) were 1 ng/mL for Sph and 0.1 ng/mL for S1P. The intra-batch and inter-batch precision was less than 15% for all quality control samples. The recoveries of the method were found to be 76.36-89.84%. The method was applied to simultaneously determine the Sph and S1P levels in mouse kidney, human plasma, and HEK 293 cells treated with tumor necrosis factor-α (TNF-α) and N,N-dimethylsphingosine (DMS). The S1P levels increased in cells treated with TNF-α whereas decreased in cells treated with DMS. These results indicated that this new LC-MS/MS method was rapid, sensitive, specific and reliable to quantify Sph and S1P levels in biological samples simultaneously. 

Sphingosine; Sphingosine 1-phosphate; C17-sphingosine; C17-sphingosine 1-phosphate; Simultaneous determination; Liquid chromatography-tandem mass spectrometry (LC-MS/MS)