Column Purchasing Caveats
SEC users should also keep in mind that some SEC packing material will slowly degrade with time. In the case of polymeric
packings stored in nonstabilized tetrahydrofuran, peroxide buildup will degrade the packing over time. Even if stabilized
solvents are used for column storage (that is, solvents that contain an antioxidant such as BHT), oxidized products might
adsorb onto the packing, changing its adsorptive properties. From extensive studies involving silica packings, long-term storage
in aqueous solutions, especially if neutral or slightly basic, can degrade silica, thereby lowering column efficiency and
generating fines that will eventually interfere with on-line light scattering detection. In view of these potential problems,
buyers should insist on purchasing newly packed columns. Although some manufacturers supply a dated test chromatogram with
each column sold, it would be more desirable for all column producers to list the manufacturing date, or specifically the
date the column was packed. This information can sometimes be traced if the column serial number is provided.
Our last caveat involving column selection is about the risk of purchasing packings that may contain inadvertently adsorbed
components on the surface. Depending on how the packing is polymerized, packed into columns, and washed, it may contain any
number of adsorbed components that could possibly change the adsorptive properties of the polymer packing. Packings can also
adsorb impurities from mobile phases or from injected samples overtime. In light of this potential problem, we recommend that
SEC columns in question be evaluated by injecting polymer samples of interest and measuring column recovery, which should
be greater than 95%. Columns also should be tested for unwanted adsorption by injecting the monomer or dimer forms of the
sample in question, and measuring elution volume or SEC distribution coefficient, K
SEC (equation 2). The K
SEC values should be close to unity to ensure the absence of adsorption. If adsorption is suspected, higher-molecular-weight samples
will probably be retained on the column, that is, K
SEC > 1, invalidating the SEC process. In the likelihood of adsorption, columns should be cleaned of adsorbed components by following
the manufacturer's recommendations, or another column brand should be tested.
Anticipated and Recommended Future Trends
SEC is a well-established technique for routine polymer and biopolymer separations, common to most laboratories that deal
with macromolecules. As such, developments in column technology have matured and slowed in recent years because many of the
parameters that affect the performance of a size-exclusion separation have been researched and optimized.
SEC separations rely heavily on the physical properties of packings, as summarized earlier (1). Thus, in this section we focus
on these inherent properties that can only be adjusted and controlled through manufacturing.
The most important packing properties are the pore size, which define the molecular weight separation range, and the pore
, or the ratio V
, which helps to establish the resolution of a separation. It appears that we are close to the maximum limit of V
of commercially available cross-linked polymer packings; to obtain a significantly higher V
ratio, a paradigm shift in column technology is required. An open network of structurally sound pores needs to be developed;
perhaps a new type of silicate needs to be realized, as with Waters BEH column technology (9).
To substantially increase V
, large-diameter columns can be used, a course taken by Polymer Standards Service's asymmetric column. An expensive proposition
is to use preparative SEC columns for analytical work. Experimentally, V
can be increased by simply adding more columns in series. In both of these cases, however, analysis time is compromised unless
higher flow rates are used.