EFFECT OF BED COMPRESSION ON HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY COLUMNS WITH GIGAPOROUS POLYMERIC PACKINGS

The behavior of chromatographic columns packed with gigaporous, highly cross-linked styrenic particles was investigated for use in protein s eparation by reversed-phase chromatography at high flow velocities. St ainless-steel columns which were 3.5 or 7.5 cm long and had an inner d iameter of 0.46 mm were slurry packed with 8 or 20 mm diameter spheric al particles of 4000 Angstrom mean pore size by using methanol as the packing fluid. It was found that the conditions employed during the pa cking process have a dramatic effect on the properties of such columns and that this can be attributed in part to the deformability of the p articles. An increase in the packing pressure to approximately 6000 p. s.i. (41 MPa) resulted in a higher mass-transfer efficiency for the co lumn with a concomitant decrease in permeability. This is ascribed to a decrease in the interstitial porosity with increasing packing pressu re since the experimentally measured plate heights for these columns w ere found to agree quantitatively with theoretical predictions that re late changes in the interstitial porosity to intraparticle mass transf er. However, the theoretically derived relationship between porosity, permeability, and efficiency does not hold for columns packed at press ures higher than 6000 p.s.i., in which case the total column porosity was found to be high while the permeability and column efficiency were low. This behavior is explained by the formation of a low-porosity la yer of highly compressed particles at the downstream end of the column during high pressure packing so that the assumption of axially unifor m column properties used in the theoretical approach leads to very lar ge errors.