# Combining Liquid Chromatography and Cryogenic IR Spectroscopy in Real Time for the Analysis of Oligosaccharides

## TL;DR

This paper introduces a real-time method combining liquid chromatography and cryogenic IR spectroscopy to better identify oligosaccharide isomers.

## Contribution

The novel approach enables real-time coupling of LC with cryogenic IR spectroscopy, overcoming previous temporal limitations.

## Key findings

- Cryogenic IR spectroscopy can be coupled with LC in real time, acquiring spectra in as little as 10 seconds.
- The method successfully identified both specified and unspecified components in a human milk oligosaccharide product.
- This approach provides an orthogonal data dimension for improved molecular identification of isomers.

## 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.