Liquid chromatography- Introduction

Liquid chromatography (LC) has become the analytical technique of reference in many fields requiring the separation and quantification of compounds present in different matrices (environmental, biological, chemical, etc.). This technique is based on the difference in distribution of compounds between two immiscible phases: a mobile phase consisting of a solvent and a stationary phase contained in a column. The interactions involved, which must be fast-acting and reversible, can be of various kinds, such as adsorption, partitioning or ion exchange. At the exit of the chromatographic column, the separated compounds are revealed on-line by means of an appropriate detector. Numerous detectors can be used, and their choice depends on the nature of the compounds separated and the sensitivities required. The two most widely used detectors today are the UV-visible spectrophotometer and the mass spectrometer. The latter has revolutionized the use of LC since the beginning of the 21st century, thanks to its extraordinary properties in terms of sensitivity, near-universality, selectivity and the structural information that can be obtained. Liquid chromatography has now largely replaced gas chromatography, which nevertheless remains the technique of choice for the analysis of volatile compounds in various fields, such as petroleum products.

From the 21st century onwards, liquid chromatography has also undergone a major evolution in terms of instrumentation, with pump systems capable of delivering the mobile phase at very high pressures (up to 1,500 bar today), very small extra-column volumes and small stationary phases enabling very rapid separations (less than a minute) and/or with very high peak capacities. This technology, known as UHPLC (Ultra High Performance Liquid Chromatography), meets the requirements of the authorities in terms of speed, sensitivity and cost of analysis, and is now widespread in fields ranging from the analysis of biological fluids to that of environmental matrices.

This article reviews this new technology, discussing the nature of stationary phases, improved instrumentation, the types of separation developed for new applications, and data processing in terms of method optimization software and validation criteria.