Comprehensive LC-MS Sphingolipid Profiling

For LC-MS analysis, sera were randomly sub-pooled into 4 groups of men and 4 groups of women, homogeneous for biometric parameters and HPTLC quantitative profiles. SL extracts were analyzed in the presence of internal standard^{52 (link)}, prepared as described by Merrill et al.^{59 (link)}, including an alkaline hydrolysis step to remove phospholipids, and analyzed by Waters Aquity Ultra Performance Liquid Chromatography (UPLC) system connected to a Waters LCT Premier orthogonal accelerated time of flight mass spectrometer (Waters, Millford, MA), operating in positive electrospray ionization mode. Spectra were acquired according to^{52 (link)}. Positive identification of compounds was based on the accurate mass measurement with an error <5 ppm and its LC retention time, compared to that of a standard (±2%). Mass spectra were analyzed by MassLynx™ 4.1 Software.
LC-MS/MS analyses of sphingosine, S1P and dhS1P were carried out on an Acquity UPLC system coupled with a Xevo TQ-MS triple quadrupole mass spectrometer. The mass spectrometer was operated in the positive ESI mode, and analytes were quantified by multiple reaction monitoring (MRM). Transitions considered were Q1 300.4- > Q3 264.4 for Sph, Q1 380.4 - > Q3 264.4 for S1P and Q1 382.4- > Q3 284.4 for dhS1P.

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Al-Daghri N.M., Torretta E., Barbacini P., Asare H., Ricci C., Capitanio D., Rosa Guerini F., Sabico S.B., Alokail M.S., Clerici M, & Gelfi C. (2019). Sphingolipid serum profiling in vitamin D deficient and dyslipidemic obese dimorphic adults. Scientific Reports, 9, 16664.

Publication 2019

Aquity Ultra Performance Liquid Chromatography (UPLC) system LCT Premier orthogonal accelerated time of flight mass spectrometer

Hydrolysis Liquid chromatography Mass spectra Ms ms Phospholipids Retention Sera Sphingosine Women

Corresponding Organization :

Other organizations : King Saud University, University of Milan, Institute of Biomedical Technologies, Don Carlo Gnocchi Foundation, Istituto Ortopedico Galeazzi, Istituti di Ricovero e Cura a Carattere Scientifico

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Variable analysis

independent variables

Sub-pooling of sera into 4 groups of men and 4 groups of women

dependent variables

LC-MS analysis of SL extracts
Quantification of sphingosine, S1P, and dhS1P by LC-MS/MS

control variables

Biometric parameters
HPTLC quantitative profiles
Internal standard for LC-MS analysis
Alkaline hydrolysis step to remove phospholipids

controls

Internal standard for LC-MS analysis

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