A Chromatographic Comparison of
Silica-C18 HPLC Columns
Introduction
Silica-based C18 columns are the most commonly used columns for
HPLC. The number of available silica-based C18 columns has risen
greatly in the past few years and now stands at more than 150. The
proliferation of C18 columns has made it difficult to choose the right
column for an application or as an appropriate backup column.
Typically chromatographers choose HPLC columns by comparing the
packing media specifications supplied by the manufacturer (i.e. surface
area, particle size and shape, carbon load, endcapping, pore diameter,
pore volume, bonding density, bonding type, etc.). This type of
comparison, although sometimes useful, can not be used to accurately
predict a column’s performance. Column selectivity and peak shape are
largely influenced by the underlying silica, and to a lesser extent, the
bonding procedure used rather than the packing material’s physical
characteristics. Therefore, the best way to compare columns is by their
chromatographic performance.
In 1995, Steffeck and co-workers1 developed a chromatographic test
and graphed the results in order to simplify comparisons of silica-based
C18 and C8 HPLC columns based on their retention, symmetry, and
selectivity for polar and non-polar compounds. The test mix used in
that work contained acidic, basic, and neutral probes. In this study, we
used a similar approach using a different test mix, and we ran the tests
at two different pH values (pH 2.5 and pH 7). Additionally, we used a
test mixture containing polyaromatic hydrocarbons to assess the
columns’ shape selectivity as described by Sander and Wise2.
To compare column performance for a wide variety of compounds under
both acidic (pH 2.5) and neutral (pH 7.0) conditions we used the test
mixture shown in Figure 1. It includes polar and non-polar probes that
are acidic, basic, or neutral. Each probe has a specific purpose in the
test mix. Uracil is used as a void-volume marker. 3-Butylpyridine is