Comparison of concentration pulse and tracer pulse chromatography: experimental determination of eluent uptake by bridged-ethylene hybrid ultra-high performance liquid chromatography packings.

Excess volume isotherms of acetonitrile and methanol sorbed on a C(18) BEH UHPLC packing were determined over a range of pressure, temperature, flow rate and eluent composition. The isotherm measurements were carried out by two independent experimental methods, viz., concentration pulse and tracer pulse chromatographies. Isotherms were measured with both experimental techniques at 30, 45 and 60 °C. The excess isotherms increased with decreasing temperature although the variations were relatively small. Direct comparison of the two experimental techniques showed that the measured void volumes were identical within experimental error. The measured excess volumes by both techniques were comparable with the concentration pulse experiments producing slightly higher excess volume data with highly aqueous eluents. Both experimental techniques show some variations of the retention volumes with sample volume, sample composition, flow rate and column inlet pressure. The results confirmed the validity of both concentration and tracer pulse chromatographies for the determination of column void volumes and the excess volume of eluent taken up by UHPLC packings.