Analysis of peptides and conjugates by amino acid analysis

Abstract

Amino acid analysis is a highly accurate method for characterization of the composition of syntheticpeptides. Together with mass spectrometry, it gives a reliable control of peptide quality and quantity beforeconjugation and immunization.Peptides are hydrolyzed, preferably in gas phase, with 6 M HCl at 110 °C for 20-24 h and the resultingamino acids analyzed by ion-exchange chromatography with post-column ninhydrin derivatization.Depending on the hydrolysis conditions, tryptophan is destroyed, and cysteine also, unless derivatized, andthe amides, glutamine and asparagine, are deamidated to glutamic acid and aspartic acid, respectively.Three different ways of calculating results are suggested, and taking the above limitations into account, aquantitation better than 5% can usually be obtained.