ABSTRACT
This chapter describes the design and application of a mixture of synthetic peptide high-performance liquid chromatography standards for monitoring free silanols on silica-based reversed-phase packings, demonstrating the versatility of the standards in monitoring silanol activity on commercial columns and columns prepared in the laboratory. The impressive sensitivity of the peptide standards, particularly Peptides 3 and 4, toward non-ideal behavior on even maximally derivatized silica supports should prove of great value to researchers involved in reversed-phase separations of peptides and other polar solutes. A deterioration in column performance also becomes apparent after extensive use of a silica-based reversed-phase column, due to the gradual removal of hydrophobic ligands from the silica, resulting in an increased concentration of underivatized silanols. Researchers utilizing reversed-phase chromatography for peptide separations would likely prefer not only to assess the level of silanol activity on a particular reversed-phase column, but also gauge the effect this activity may have on the retention behavior of the peptide solutes.
