A Hyphenated Technique based on High-Performance Thin Layer Chromatography for Determining Neutral Sphingolipids: A Proof of Concept

Chromatography

Andrés Domínguez¹  Carmen Jarne¹  Vicente L. Cebolla¹  Javier Galbán⁴  María Savirón²  Jesús Orduna⁵  Luis Membrado¹  María-Pilar Lapieza¹  Elena Romero¹  Isabel Sanz Vicente⁴  Susana de Marcos⁴  Rosa Garriga³ 

[1] ICB-CSIC, C/ Miguel Luesma, 4, 50018 Zaragoza, Spain; E-Mails:;CEQMA-CSIC, Facultad de Ciencias, Universidad de Zaragoza, 50009 Zaragoza, Spain; E-Mail:;Departamento de Química-Física, Universidad de Zaragoza, 50009 Zaragoza, Spain; E-Mail:;Departamento de Química Analítica, Universidad de Zaragoza, 50009 Zaragoza, Spain; E-Mails:;ICMA-CSIC, Facultad de Ciencias, Universidad de Zaragoza, 50009 Zaragoza, Spain; E-Mail:

关键词: TLC-MS;    induced fluorescence;    automated multiple development;    primuline;    sphingolipids;   

DOI  :  10.3390/chromatography2020167

来源: mdpi

【 摘 要 】

Hyphenated HPTLC has been used to analyze several neutral sphingolipids acting as lysosomal storage disease (LSD) biomarkers. Automated multiple development (AMD) provides separation of lipid peaks, which are detected and quantified using fluorescence detection by intensity changes (FDIC) after primuline post-impregnation. A final online transfer to a mass spectrometer by means of an elution-based interface allows their identification using electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI).Given that the increases in fluorescent emission detected by FDIC are produced by non-specific, electrostatic interactions between the primuline and hydrocarbon chains in the ceramide backbones of sphingolipids, it is a non-destructive detection technique, allowing the precise location and transfer of biomarker peaks to a mass spectrometer using an elution interface. By using primuline as a fluorophore, the technique is also compatible with ESI-APCI and does not interfere with the MS of sphingolipids. APCI provides useful and complementary structural information to the ESI for sphingolipid identification. Moreover, FDIC emission can be used for quantitative purposes. Results include the determination of sphingomyelin (SM) in human-plasma samples (RSD < 6%) by means of a standard addition method with non-linear calibration, and the identification of globotriaosylceramide (Gb3) in the plasma of a Fabry patient. Only one HPTLC plate is needed to perform the analysis.

【 授权许可】

CC BY   
© 2015 by the authors; licensee MDPI, Basel, Switzerland.