Analysis of Gangliosides Directly from Thin-Layer Chromatography Plates by Infrared Matrix-Assisted Laser Desorption/Ionization Orthogonal Time-of-Flight Mass Spectrometry with a Glycerol Matrix

• Published in: Analytical chemistry (Washington) Vol. 77; no. 13; pp. 4098 - 4107
• Main Authors: Dreisewerd, Klaus, Müthing, Johannes, Rohlfing, Andreas, Meisen, Iris, Vukelić, Željka, Peter-Katalinić, Jasna, Hillenkamp, Franz, Berkenkamp, Stefan
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 01.07.2005
• Subjects: Animals; CHO Cells; Chromatography; Chromatography, Thin Layer - methods; Cricetinae; Cricetulus; Dyes; G(M3) Ganglioside - chemistry; G(M3) Ganglioside - metabolism; Glycerol - chemistry; Lasers; Mass spectrometry; Sensitivity; Silica; Spectrometry, Mass, Electrospray Ionization - methods; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization - methods
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac048373w

A novel method is presented for direct coupling of high-performance thin-layer chromatography (HPTLC) with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) for the analysis of biomolecules. A first key feature is the use of a liquid matrix (glycerol), which provides a homogeneous wetting of the silica gel and a simple and fast MALDI preparation protocol. A second is the use of an Er:YAG infrared laser, which ablates layers of ∼10-μm thickness of analyte-loaded silica gel and provides a soft desorption/ionization of even very labile analyte molecules. The orthogonal time-of-flight mass spectrometer employed in this study, finally provides a high accuracy of the mass determination, which is independent of any irregularity of the silica gel surface. The analytical potential of the method is demonstrated by the compositional mapping of a native GM3 (II3-α-Neu5Ac−LacCer) ganglioside mixture from cultured Chinese hamster ovary cells. The analysis is characterized by a high relative sensitivity, allowing the simultaneous detection of various major and minor GM3 species directly from individual HPTLC analyte bands. The lateral resolution of the direct HPTLC-MALDI-MS analysis is defined by the laser focus diameter of currently ∼200 μm. This allows one to determine mobility profiles of individual species with a higher resolution than by reading off the chromatogram by optical absorption. The fluorescent dye primuline was, furthermore, successfully tested as a nondestructive, MALDI-compatible staining agent.