ABSTRACT
This chapter describes the design and application of a series of synthetic peptide size-exclusion standards and demonstrates their versatility in monitoring both ideal and non-ideal size-exclusion chromatography (SEC) column performance for peptide separations. It also describes three size-exclusion columns: Altex Spherogel TSK G2000SW, SynChropak GPC60, and Pharmacia Superose 12. The synthetic peptide were designed specifically for assessing the peptide resolving power of size-exclusion columns and are capable of monitoring both ideal and non-ideal chromatographic behavior. The ideal size-exclusion behavior of the peptide standards was obtained with a volatile 0.1% aqeous trifluoroacetic acid (TFA) eluent. The ability to predict the position and/or elution order of peptides during SEC of a peptide mixture greatly simplifies preliminary isolation of peptides from a chemical or proteolytic protein digest. The adoption of a common set of peptide standards by chromatographers utilizing SEC could greatly facilitate an accurate comparison of results between different researchers.
