MICROCOLUMN SIZE-EXCLUSION CHROMATOGRAPHY WITH POLYMERIC STATIONARY PHASES

Miniaturization of the column used in liquid chromatographic separations to microcolumn dimensions (0.1-0.5-mm i.d.) offers various advantages to the technique, such as low eluent consumption, ability to interface to other chromatographic techniques and mass spectrometers, low cost per column, and reduced maintenance requirements, among others. This report documents studies conducted in the preparation of highly efficient microcolumns for size exclusion chromatography (micro SEC) with polymer packings of 50- and 1000-angstrom pore size. Studies conducted on the effects of slurry ratio and column packing pressure on efficiency indicate that higher packing pressures yield the lowest plate heights (higher efficiency) but at the expense of decreased specific column permeabilities, while slurry ratio only has a significant effect on plate height at low packing pressures (170 atm). Results obtained suggest that short columns (30 cm) should be prepared at 400 atm with high slurry concentrations while long columns (1 m) are best prepared at low packing pressures with dilute slurries. Columns with approximately 75 000 plates/m can be routinely prepared. Resolution factors obtained (Dsigma) were 0.033 for a conventional size column, 0.017 for 1-m microcolumns operated at the optimum linear velocity; and 0.021 for the microcolumns operated at a linear velocity chosen to yield retention times equivalent to the conventional size column. The results obtained indicate better performance for the micro SEC system when compared to conventional size SEC.