Ganglioside analysis by thin-layer chromatography matrix-assisted laser desorption/ionization orthogonal time-of-flight mass spectrometry

Abstract

Metastable decomposition of ions generated in matrix-assisted laser desorption/ionization (MALDI) mass spectrometers complicates analysis of biological samples that have labile bonds. Recently, several academic laboratories and manufacturers of commercial instruments have designed instruments that introduce a cooling gas into the ion source during the MALDI event and have shown that the resulting vibrational cooling stabilizes these labile bonds. In this study, we compared stabilization and detection of desorbed gangliosides on a commercial orthogonal time-of-flight (oTOF) instrument with results we reported previously that had been obtained on a home-built Fourier transform mass spectrometer. Decoupling of the desorption/ ionization from the detection steps resulted in an opportunity for desorbing thin-layer chromatography (TLC)-separated gangliosides directly from a TLC plate without compromising mass spectral accuracy and resolution of the ganglioside analysis, thus coupling TLC and oTOF mass spectrometry. The application of a declustering potential allowed control of the matrix cluster and matrix adduct formation, and, thus, enhanced the detection of the gangliosides. during matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) analyses of molecules having labile bonds. Although this phenomenon can be used for obtaining analyte structural information for small peptides, oligosaccharides, and some glycolipids [1-5] for larger or more fragile molecules, it can limit the mass accuracy and resolution and complicate interpretation of the spectra of heterogeneous samples. Thus, it is important to develop methods that allow control of the extent of metastable decay. In addition to the pressure of the background gas and extraction field strength in the MALDI technique, the major factors affecting metastable fragmentation are laser wavelength/pulse width and the choice of the MALDI matrix [6,7]. In addition, the presence of electrons may play a role in analyte fragmentation and may cause suppression of multiply charged ions in a spectrum [8,9].Control of the metastable fragmentation in MALDI by increasing the pressure in the ionization source and reduction of the ions internal energy was first introduced by Loboda et al. on orthogonal time-of-flight (oTOF) instruments [10] and later extended to Fourier transform mass spectrometry (FTMS) [11] and to the operation at atmospheric pressure on various types of instruments [12]. These methods have since been used for stabilization of glycolipids [13,14] and peptides and proteins [11,15]. The term vibrational cooling (VC) MALDI recently has been introduced to describe desorption in the pressure range where cooling of the excess bond energy is achieved [12]. Stabilization of the labile bonds under these conditions allows for control of the extent of fragmentation, which is particularly

Analysis of native milk oligosaccharides directly from thin-layer chromatography plates by matrix-assisted laser desorption/ionization orthogonal-time-of-flight mass spectrometry with a glycerol matrix

“…The detection limit in the mass range below 1000 u is apparently mostly determined by the background from the silica gel-matrix system. [18,32]. Besides for the isoform distinction, an unambiguous identification of the oligosaccharide species composition can be achieved.…”

High-sensitivity analysis of glycosphingolipids by matrix-assisted laser desorption/ionization quadrupole ion trap time-of-flight imaging mass spectrometry on transfer membranes

“…This method largely eliminates the disadvantages of the indirect method, thus increasing the utility of TLC for structural characterization [28][29][30][31][32][33]. Direct TLC-MS presents difficulties with regard to complete retrieval of the target compound from the silica coated on the plate and could also cause contamination of silica to the MS instrument.…”

“…Recently, remarkable advances achieved in the development of MALDI MS ion trap instruments have enabled TLC-MALDI MS analysis to be performed with high mass accuracy and sensitivity despite the irregular surface of the plate [35]. Some instruments include an external ion trap, such as a Fourier transform ion cyclotron resonance ion trap (FTICR) [28,29] and an orthogonal extracting ion trap [30,31] could also apply to TLC-MALDI MS. Developed GM1 was difficult to be identified even by using highly sensitive TLC-FTICR MS at the amount of 120 fmol [28].…”

“…The aim of the previous study was to analyze the visible GSLs with the conventional staining method; i.e., GSLs were analyzed based on the combined information from TLC and MS. Due to the similarity of the properties of the procedures, the limits of detection of these studies were the same as that of the conventional staining method; 10 pmol [30,31]. However, we tried to detect far more minor components to fully identify the components of the sample, regardless of the detection limit of staining, by the use of the imaging mass spectrometry method.…”

Vacuum compatible sample positioning device for matrix assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry imaging

“…Although 0.1 μm positioning accuracy might seem somewhat excessive for the purposes, recent developments in laser optics within mass spectrometry community [12][13][14][15][16] allows acquiring MALDI with submicron laser spot diameters. Clearly, for such small areas, laser power must be significantly increased causing excessive fragmentation, yet vibrational cooling [32][33][34][35][36][37][38] should prove itself useful to reduce such fragmentation.…”