Influence of pressure on the properties of chromatographic columns

The effect of the local pressure and of the average column pressure on the hold-up column volume was investigated between 1 and 400 bar, from a theoretical and an experimental point of view. Calculations based upon the elasticity of the solids involved (column wall and packing material) and the compressibility of the liquid phase show that the increase of the column hold-up volume with increasing pressure that is observed is correlated with (in order of decreasing importance): (1) the compressibility of the mobile phase (+1 to 5%); (2) in RPLC, the compressibility of the C18-bonded layer on the surface of the silica (+0.5 to 1%); and (3) the expansion of the column tube (< 0.001%). These predictions agree well with the results of experimental measurements that were performed on columns packed with the pure Resolve silica (0% carbon), the derivatized Resolve-C18 (10% carbon) and the Symmetry-C18 (20% carbon) adsorbents, using water, methanol, or n-pentane as the mobile phase. These solvents have different compressibilities. However, 1% of the relative increase of the column hold-up volume that was observed when the pressure was raised is not accounted for by the compressibilities of either the solvent or the C18-bonded phase. It is due to the influence of the pressure on the retention behavior of thiourea, the compound used as tracer to measure the hold-up volume.