Wheat gluten amino acid composition analysis by high-performance anion-exchange chromatography with integrated pulsed amperometric detection

• Published in: Journal of Chromatography A 2009-07, Vol.1216 (29), p.5557-5562
• Main Authors: Rombouts, Ine, Lamberts, Lieve, Celus, Inge, Lagrain, Bert, Brijs, Kristof, Delcour, Jan A.
• Format: Article
• Language: English
• Subjects: acid treatment; Amino acid; amino acid composition; Biological and medical sciences; Cereal and baking product industries; electrochemistry; Food industries; Fundamental and applied biological sciences. Psychology; gliadin; Gluten; glutenins; high performance anion exchange chromatography; High-performance anion-exchange chromatography with integrated pulsed amperometric detection; HPAEC-IPAD; hydrochloric acid; integrated pulsed amperometric detection; ion exchange chromatography; pretreatment; Wheat; wheat gluten
• ISSN: 0021-9673
• DOI: 10.1016/j.chroma.2009.05.066

A simple accurate method for determining amino acid composition of wheat gluten proteins and their gliadin and glutenin fractions using high-performance anion-exchange chromatography with integrated pulsed amperometric detection is described. In contrast to most conventional methods, the analysis requires neither pre- or post-column derivatization, nor oxidation of the sample. It consists of hydrolysis (6.0 M hydrochloric acid solution at 110 °C for 24 h), evaporation of hydrolyzates (110 °C), and chromatographic separation of the liberated amino acids. Correction factors ( f) accounted for incomplete cleavage of peptide bonds involving Val ( f = 1.07) and Ile ( f = 1.13) after hydrolysis for 24 h and for Ser ( f = 1.32) losses during evaporation. Gradient conditions including an extra eluent (0.1 M acetic acid solution) allowed multiple sequential sample analyses without risk of Glu contamination on the anion-exchange column. While gluten amino acid compositions by the present method were mostly comparable to those obtained by a conventional method involving oxidation, acid hydrolysis and post-column ninhydrin derivatization, the latter method underestimated Tyr, Val and Ile levels. Results for the other amino acids obtained by the different methods were linearly correlated ( r > 0.99, slope = 1.03).