Handbook of Size Exclusion Chromatography and Related ...

Table 1 Comparison Between SEC and HdC Separations
shown in Fig. 5, is slightly narrower than that of asingle-pore-size SEC column.
Moreimportantly,itbecomesmoreandmoredifficulttouseacolumnpackedwith
particles smaller than 1.0mm due to the increase in backpressure. The separation
of low MW species by HdC quickly diminishes below 10 4 MW.
The SEC separation mechanism of high MW polymers, which involves an
in-and-out-of-pore process, becomes more difficult for high MW polymers. The
higher the MW,the slower the movement, and the more difficult the separation.
ThehighMWpolymerchainsarealsomoresusceptibletodegradationduringthis
in-and-out-of-pore process. Therefore, the peak shape of high MW standards,
.10 6 ,separated by large-pore columns tends to be broader and exhibits tailing.
Ontheotherhand,theseparationmechanismofHdCdoesnotinvolvethisin-andout-of-poreprocess.ThisisthereasonthatthehighMWpeaksinFig.1,separated
by small pore columns, tends to be sharp.
These two separation mechanisms co-exist in achromatographic run and
complementeachother,asshowninTable1.Itwouldbeidealtodesignacolumn
thatusestheadvantagesofeachmechanismandprovidesalinearcalibrationcurve.
4 COMBINATION OF SEC AND HdC
SEC HdC
Location where separation Pores inside gels Interstitial area between
occurs
gels
Factors affecting separation Pore size & pore volume Particle size
For low MW material MW range & slope of
calibration curve can be
easily designed
For high MW material Poorer separation due to
slow process, and
possible degradation
Separation diminishes
below 10 4 MW
More favorable
Currently,HdC studies for MW separations emphasize the same MW ranges as
regular SEC studies (2). The particle size of HdC packings is normally less than
3mm. In order to separate polymers in the higher MW range for the SEC/HdC
combination, the particle size should be larger.Three columnsare custom-packed
with 3, 5, and 10mm solid beads by Jordi’sAssociate (Bellingham, MA, USA).
The calibration curves of polystyrene standards in THF are shown in Fig. 8. All
three columns appear to have an inflection point around 5 10 5 . The curves are
steeper below this point, and are less steep above it. The slopes at the upper MW
© 2004 by Marcel Dekker, Inc.