Size-exclusion high-performance liquid chromatography of peptides: a review

Gel filtration, now referred to as size exclusion chromatography (SEC), on soft gels has been employed for over 40 years for the separation, desalting and molecular weight estimation of peptides and proteins. Developments in rigid particle technology during the 1970s led to the commercial availability of pre-packed columns giving rapid run times and increased resolution, referred to as size-exclusion high-performance chromatography (SE-HPLC). Initially these packings were more suitable for the separation of proteins than of peptides. Today, supports that operate in the fractionation range <10000 Daltons (Da) are available. In this review, the theoretical basis of SEC is discussed and examples are given of its application to the study of peptides, especially regarding purification, estimation of molecular weight and study of molecular associations.