Sphingolipids have hydrophilic and hydrophobic properties, different saturation and combination of the oligosaccharide chains and mass homology of species located in a narrow m/z region hampering their recognition. To target sphingolipids for diagnostic purposes, standardized methods for lipid extraction, quali- and quantitative assessments are required. In this study, HPTLC-MALDI MS was adopted to establish sphingolipid and glycosphingolipid profiles in muscle, brain and serum to create a database of molecules to be searched in the preclinical and clinical investigations. Specific protocols for lipid extraction were set up based on the characteristics of the tissue or/and fluids; this approach maximizes the HPTLC-MALDI MS analytical throughput both for lipids extracted in organic and aqueous phase. This study indicates that alkaline hydrolysis is necessary for the detection of low abundant species such as Gb3Cer and ceramides in serum and Gb4Cer, CerP and HexCer in muscle tissue. The high hydrophobicity of ceramides has been overcome by the development of HPTLC plate in chloroform:methanol/50:3.5, which increases the number and the intensity of low abundant Cer species. MS/MS analysis has been conducted directly on HPTLC plate allowing the molecular recognition; furthermore a dataset of spectra was acquired to create a database for future profiling of these molecules.

Torretta, E., Fania, C., Vasso, M., Gelfi, C. (2016). HPTLC-MALDI MS for (glyco)sphingolipid multiplexing in tissues and blood: A promising strategy for biomarker discovery and clinical applications. ELECTROPHORESIS, 37(14), 2036-2049 [10.1002/elps.201600094].

HPTLC-MALDI MS for (glyco)sphingolipid multiplexing in tissues and blood: A promising strategy for biomarker discovery and clinical applications

Fania, Chiara;
2016

Abstract

Sphingolipids have hydrophilic and hydrophobic properties, different saturation and combination of the oligosaccharide chains and mass homology of species located in a narrow m/z region hampering their recognition. To target sphingolipids for diagnostic purposes, standardized methods for lipid extraction, quali- and quantitative assessments are required. In this study, HPTLC-MALDI MS was adopted to establish sphingolipid and glycosphingolipid profiles in muscle, brain and serum to create a database of molecules to be searched in the preclinical and clinical investigations. Specific protocols for lipid extraction were set up based on the characteristics of the tissue or/and fluids; this approach maximizes the HPTLC-MALDI MS analytical throughput both for lipids extracted in organic and aqueous phase. This study indicates that alkaline hydrolysis is necessary for the detection of low abundant species such as Gb3Cer and ceramides in serum and Gb4Cer, CerP and HexCer in muscle tissue. The high hydrophobicity of ceramides has been overcome by the development of HPTLC plate in chloroform:methanol/50:3.5, which increases the number and the intensity of low abundant Cer species. MS/MS analysis has been conducted directly on HPTLC plate allowing the molecular recognition; furthermore a dataset of spectra was acquired to create a database for future profiling of these molecules.
Articolo in rivista - Articolo scientifico
Gangliosides; Glycosphingolipids; HPTLC-MALDI MS; Multiplexing; Sphingolipids;
Gangliosides; Glycosphingolipids; HPTLC-MALDI MS; Multiplexing; Sphingolipids; Animals; Biomarkers; Brain; Chromatography, Thin Layer; Glycosphingolipids; Hydrolysis; Mice; Mice, Inbred C57BL; Muscle, Skeletal; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
English
2016
37
14
2036
2049
none
Torretta, E., Fania, C., Vasso, M., Gelfi, C. (2016). HPTLC-MALDI MS for (glyco)sphingolipid multiplexing in tissues and blood: A promising strategy for biomarker discovery and clinical applications. ELECTROPHORESIS, 37(14), 2036-2049 [10.1002/elps.201600094].
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/10281/215243
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