Structure characterisation using thin-layer chromatography-MALDI MS and infrared ion spectroscopy

The integration of thin-layer chromatography (TLC) with matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) provides a fast workflow for the analysis of mixtures. Nevertheless, distinguishing isomers and the structural elucidation of unknown compounds remains challenging and resource intensive. Based on tandem mass spectrometry, infrared ion spectroscopy (IRIS) generates structurally diagnostic vibrational spectra for mass-selected ions confined within a mass spectrometer. By coupling IRIS to the TLC-MALDI-MS workflow, the IR spectra of molecules ionized directly from the TLC plate are obtained. In this study, we separated and identified three isomeric compounds in a mixture through sequential TLC separation, ionization, mass selection and characterization using IRIS. Furthermore, we demonstrate isomer-selective photofragmentation scans of the TLC plate along the axis of separation. This TLC-MALDI-IRIS approach represents a robust, sensitive, and versatile analytical method with broad applications in complex mixture analysis. • Thin-layer chromatography (TLC), matrix-assisted laser desorption ionization (MALDI), high-resolution mass spectrometry (MS) and infrared ion spectroscopy (IRIS) have been integrated in a single workflow. • Proof-of-concept measurements were conducted using three closely similar isomeric compounds, separated by TLC and characterized by IRIS. • The TLC-MALDI-IRIS results were compared to TLC-MALDI-MS/MS for structure identification giving a confidence score. • Single-frequency isomer-selective scans were measured from the TLC plate demonstrating a higher-throughput approach for isomer distinction.