Two-dimensional liquid chromatography (2D-LC)

Liquid chromatography (LC) is the analytical technique of choice in many fields (see [P 1 655][P 1 494] and [P 1 460] ). Indeed, the various LC modes (RPLC, IEX, SEC, HIC, HILIC, etc.) enable the separation power to be adapted to the physico-chemical properties of the molecules to be analyzed. However, the growing need for fine characterization of complex media, combined with increasingly stringent regulatory requirements, calls for a separation power (or peak capacity) that one-dimensional LC (1D-LC) cannot achieve. This is despite the use of ultra-high pressure systems (1,200 to 1,500 bar) combined with columns equipped with particle diameters of less than 2 μm (UHPLC). Two-dimensional liquid chromatography (2D-LC) is the answer to these challenges. Indeed, 2D-LC combines two complementary separation dimensions, increasing the separation space and therefore the overall resolving power. What's more, adding a second chromatographic separation makes it easier to couple mass spectrometry to incompatible or difficult-to-compare chromatographic modes (IEX, HIC, NPLC, etc.), giving access to an additional dimension of information. The same reasoning applies to existing methods that are difficult to modify (e.g. pharmaceutical methods).

This article describes the principle of 2D-LC, the different approaches available (targeted/non-targeted, off-line/on-line) and defines the criteria for choosing its use over a 1D-LC method. Key parameters for the development of 2D-LC methods are described, focusing on the 2D-LC "comprehensive" on-line mode, which is the technique with the greatest potential, but for which method development remains complex. Finally, several examples of applications are presented in a wide range of fields.

As is customary in the industry, the percentages given are by volume, unless otherwise specified.