The optimization of the Osborne Extraction Method for the fractionation and characterization of oat proteins

Abstract

The growing number of wheat-related allergies worldwide has resulted in a new trend towards gluten-free alternatives. In this context, alternative cereals such as sorghum and oats are attracting new interest. Given the limited data available, the question of whether these cereals are completely safe and gluten-free for allergy sufferers remains open. One of the key steps in protein research is their efficient extraction. In this work, the Osborne sequential extraction method was developed and optimized using the response surface methodology in order to fractionate oat proteins. An optimized desirability of 0.986 was achieved with an extraction time of 4.7 min, a speed of 6, and a sample/solvent ratio of 5. The corresponding optimized responses were 8.7, 4.0, and 5.1% for the extraction yields of the avenin, avenalin, and albumin/globulin fractions, respectively. Further characterization of the extracts was carried out on 24 homogeneous and commercial oat samples via LC-MS/MS, targeting six potentially allergenic proteins. The avenin-E protein featured prominently, with relative contents of 60.7, 32.2, 58.0, and 59.8% in the total extract, avenin, avenalin, and albumin/globulin fractions, respectively, while the Avenin-3, ATI-2, avenin, SSG2, and SSG1 proteins in the total extract showed levels of 16.4, 9.3, 6.6, 4.8, and 2.2%, respectively. The preliminary results of an ELISA performed on the different fractions revealed low levels of gluten (from 1.24 ± 0.14 to 3.61 ± 0.16 mg/kg), which were well below the threshold limit of 20 mg/kg. These results support the hypothesis that oats can be a safe food for people suffering from cereal-related allergies. These results open the door to further studies into the comprehensive characterization of oat proteins.