Original Research ARTICLE
Preparation and characterization of avenin-enriched oat protein by chill precipitation for feeding trials in celiac disease.
- 1School of Biomedical Sciences, The University of Melbourne, Australia
- 2School of Science, Edith Cowan University, Australia
- 3School of Veterinary and Life Sciences, Murdoch University, Australia
- 4Other, Australia
- 5Walter and Eliza Hall Institute of Medical Research, Australia
- 6Department of Medical Biology, Faculty of Medicine, Dentistry and Health Sciences, University of Melbourne, Australia
- 7Department of Gastroenterology, The Royal Melbourne Hospital, Australia
- 8Murdoch Childrens Research Institute (MCRI), Australia
The safety of oats for people with celiac disease remains unresolved. While oats have attractive nutritional properties that can improve the quality and palatability of the restrictive, low fibre gluten-free diet, rigorous feeding studies to address their safety in celiac disease are needed. Assessing the oat prolamin proteins (avenins) in isolation and controlling for gluten contamination and other oat components such as fibre that can cause non-specific effects and symptoms is crucial. Further, the avenin should contain all reported immunogenic T cell epitopes, and be deliverable at a dose that enables biological responses to be correlated with clinical effects. To date, isolation of a purified food-grade avenin in sufficient quantities for feeding studies has not been feasible. Here, we report a new gluten isolation technique that enabled 2 kg of avenin to be extracted from 400 kg of wheat-free oats under rigorous gluten-free and food grade conditions. The extract consisted of 85% protein of which 93% of the protein was avenin. The concentration of starch (1.8% DW), -glucan (0.2% DW) and free sugars (1.8% DW) were all low in the final avenin preparation. Other sugars including oligosaccharides, small fructans and other complex sugars were also low at 2.8% DW. Liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis of the proteins in these preparations showed they consisted only of oat proteins and were uncontaminated by gluten containing cereals including wheat, barley or rye. Proteomic analysis of the avenin enriched samples detected more avenin subtypes and fewer other proteins compared to samples obtained using other extraction procedures. The identified proteins represented five main groups, four containing known immune-stimulatory avenin peptides. All five groups were identified in the 50% (v/v) EtOH extract however the group harbouring the epitope DQ2.5-ave-1b was less represented. The avenin-enriched protein fractions were quantitatively collected by reversed phase HPLC and analysed by MALDI-TOF MS. Three reverse phase HPLC peaks, representing approximately 40% of the protein content, were enriched in proteins containing DQ2.5-ave-1a epitope. The resultant high quality avenin will facilitate controlled and definitive feeding studies to establish the safety of oat consumption by people with celiac disease.
Keywords: Oats, Avenin, gluten-free diet, LC-MS, MALDI-TOF, Celiac Disease
Received: 31 Jul 2019;
Accepted: 26 Sep 2019.
Copyright: © 2019 Tanner, Juhász, Florides, Nye-Wood, Békés, Colgrave, Russell, Hardy and Tye-Din. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Mx. Gregory J. Tanner, School of Biomedical Sciences, The University of Melbourne, Melbourne, Australia, firstname.lastname@example.org