Handbook of Size Exclusion Chromatography and Related ...

materials are commercially available for SEC, including both silica- and polymerbased
media. This chapter discusses in detail the technology and application of
polymer-based packings for SEC using both organic- and aqueous-based eluents.
2 COLUMN PACKING AND PERFORMANCE
Columns of semirigid polymer gels are generally packed using a balanced density
slurry packing technique at pressures in the range 2000–4000 psi (3). Column
internal diameters of 7–8 mm i.d. have been employed traditionally, although in
recent years narrow bore (4–6 mm i.d.) columns have become more commonplace
for environmental and safety reasons because they require reduced solvent
consumption. Column lengths are typically 200–600 mm and the overall
dimensions of SEC columns available today represent a good compromise
between resolution and analysis time using flow rates and operating pressures in
accordance with common high-performance liquid chromatography equipment.
Column performance is usually assessed by performing a plate count
measurement using a relatively low viscosity eluent and a totally permeating test
probe, such as toluene in tetrahydrofuran for organic-based packings or glycerol in
water for aqueous SEC columns (4,5). Several methods for measuring plate count
(N) from the elution profile of the test probe are well documented and Fig. 1
illustrates the commonly used half height method for plate count calculation as
well as the symmetry factor. This type of column test is useful because it provides
reference performance data for future comparison during the lifetime of the
column. It is important to remember, however, that such data should always be
© 2004 by Marcel Dekker, Inc.
Figure 1 Calculation of plate count, N, and symmetry factor.