Liquid Chromatography

High-Performance Liquid Chromatography is a widely used technique for the separation and characterization of complex mixtures due to its versatility, and also due to its wide field of application in all branches of Science.

The components of the sample, previously treated and dissolved, are introduced into a suitable eluent (mobile phase), and by applying high pressures they are forced through a chromatographic column (stationary phase). This produces a selective retention of the analytes, and the resolution of this separation depends on the interaction of these with the stationary phase and the mobile phase. The separation can be optimized by varying the mixtures or composition of the mobile phase, as well as the nature of the column packing.

We can classify Liquid Chromatography as a second or separation method:

• Liquid, solid or adsorption: Adsorption on the surface of the stationary phase
• Liquid: Distribution between liquid phases, one mobile and one stationary
• Bound phase: Partition and/or adsorption between the mobile and bound phases
• Affinity: Use of immobilized ligand structure to bioselectively bind the desexed protein.

Once separated, the compounds are detected using a specific detector, generating a characteristic signal that depends on both the type of compound and its concentration. The graphical representation of the signal versus the retention time of the compounds is called a chromatogram.