High-pressure liquid chromatography of peptides and proteins XI. The use of cationic reagents for the analysis of peptides by high-pressure liquid chromatography

• Published in: Journal of Chromatography A 1979, Vol.168 (2), p.377-384
• Main Authors: Hancock, W.S., Bishop, C.A., Battersby, J.E., Harding, D.R.K., Hearn, M.T.W.
• Format: Article
• Language: English
• ISSN: 0021-9673
• DOI: 10.1016/0021-9673(79)80007-2

This report describes the effect of different cationic reagents (tetraalkylammonium, alkylammonium and inorganic salts) on the retention times of di- to pentapeptides chromatographed on a reversed-phase support ( i.e. a μBondapakalkylphenyl column). Several trends are apparent with these reagents which can be explained on the basis of either ion-pairing or ion-exchange interaction of the reagent with the peptide sample. Reagents which generate in solution small highly solvated cations, e.g. Li, Na or Mg salts, give retention times similar to those obtained for ammonium salts. Tetraethylammonium salts give a modest increase in retention times relative to ammonium salts. By contrast, hydrophobic cations with long or bulky carbon chains. e.g. tetrabutylammonium or dodecylammonium ions, cause substantial decreases in retention times, resulting in very rapid elution of all peptides examined from the reversed-phase column. These observations are consistent with the composite interplay of ion-pair partition and dynamic ion-exchange effects for the cationic reagents. The use of a mixture of dodecylammonium acetate and sodium dodecylsulphate for the analysis of peptides and proteins is described. It is anticipated that such a chromatographic system will he useful for the analysis of proteins which readily aggregate.