Abstract
While the combination of liquid chromatography (LC) and mass spectrometry (MS) serves as a robust approach for oligosaccharide analysis, it has difficulty distinguishing the smallest differences between isomers. The integration of infrared (IR) spectroscopy within a mass spectrometer as an additional analytical dimension can effectively address this limitation by providing a molecular fingerprint that is unique to each isomer. However, the direct interfacing of LC-MS with IR spectroscopy presents a technical challenge arising from the mismatch in the operational time scale of each method. In previous studies, this temporal incompatibility was mitigated by employing strategies designed to slow down or broaden the LC elution peaks of interest, but this workaround is applicable only for a few species at a time, necessitating multiple LC runs for comprehensive analysis. In the current work, we directly couple LC with cryogenic IR spectroscopy by acquiring a spectrum in as little as 10 s. This allows us to generate an orthogonal data dimension for molecular identification in the same amount of time that it normally takes for LC analysis. We successfully demonstrate this approach on a commercially available human milk oligosaccharide product, acquiring spectral information on the eluting peaks in real time and using it to identify both the specified constituents and nonspecified product impurities.