IR-MALDI-MS Analysis of HPTLC-Separated Phospholipid Mixtures Directly from the TLC Plate

Abstract

The application of a recently developed direct coupling of high-performance thin-layer chromatography (HPTLC) and infrared matrix-assisted laser desorption/ionization orthogonal extracting time-of-flight mass spectrometry (Dreisewerd, K.; Müthing, J.; Rohlfing, A.; Meisen, I.; Vukelic, Z.; Peter-Katalinic, J.; Hillenkamp, F.; Berkenkamp, S. Anal. Chem. 2005, 77, 4098-4107) to the analysis of phospholipid mixtures is demonstrated. Mixtures of six phospholipid types were exemplarily analyzed. The sensitivity was found to be in the range between about 10 and 150 pmol of material spotted for HPTLC, depending on phospholipid acidity, Rf value, and ion polarity. The lateral resolution of the analysis is on the order of the laser focus diameter of about 220 x 300 microm2, allowing differentiation between phospholipid species of different acyl chain composition within one single HPTLC band, which were undistiguishable by a mere visual assessment. Analyte diffusion due to the addition of glycerol to the HPTLC plate was found to be-if at all notable-of only minor importance.

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

“…There have been previous reports in which the detailed distribution of molecular species in a band of glycolipid [30] or phospholipid [38] was shown using mass spectra alone. But the present report on detection of GM1 is quite new in that such a result was visualized by the imaging data with µm-order resolution, and further that structural analyses by MS/MS and MS/ MS/MS analyses demonstrated the precise structure of these molecular species.…”

“…There have been only a few recent reports on TLC-Blot-MALDI MS, because the additional work, i.e., the transfer from TLC plates to PVDF membranes, requires, and the orthogonal configuration of the instrument allows for direct desorption of samples from the irregular surface of the TLC plate without impairment of either mass accuracy or resolution [38]. However Costello et al have mentioned in their report that the spectral quality is better on PVDF membranes than on TLC plates, in terms of sensitivity, mass resolution, and background interference [36].…”

Surface analysis of lipids by mass spectrometry: More than just imaging

“…In addition, the greater laser penetration depth means IR-MALDI is also well suited for analysis of TLC plates using a glycerol matrix, while there are also numerous reports of successful application of intermediate and atmospheric pressure sources [107,109,110,[113][114][115].…”

Phospholipid Analysis by Nanoflow Liquid Chromatography-Tandem Mass Spectrometry

“…Analysis of intact PLs without prior derivatization has been carried out with thin layer chromatography (TLC) using densitometry 15,16 or matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF-MS). 17,18 Rapid advances in MS technology have led to sophisticated detection of PLs using electrospray ionization (ESI). While ESI-MS alone allows high speed analysis of lipid molecules, 4,19,20,21 lipids with complicated structures and ion suppression from high abundant species are issues even for high-resolution MS. Coupling liquid chromatography (LC) to ESI-MS/MS 20,22,23 has facilitated the structural determination of lipid species from complicated mixtures, enabling generation of extensive libraries of lipids.…”