Detection of gluten in a pilot-scale barley-based beer produced with and without a prolyl endopeptidase enzyme

Immunochemical and mass spectrometric methods were used to examine the gluten composition of a gluten-reduced beer produced by brewing with barley malt in the presence of prolyl endopeptidase (PEP) and a final filtration treatment with diatomaceous earth and perlite. The competitive ELISA is generally considered appropriate for the analysis of hydrolysed gluten, but it is not considered a scientifically valid method for the quantification of gluten in fermented or hydrolysed foods due to the lack of an appropriate reference standard. As no single analytical method can capture the spectrum of gluten-derived products in beer, a comprehensive approach was employed to analyse the intact and hydrolysed fractions of gluten with complementary methods. The combination of PEP addition and diatomaceous earth/perlite filtration was more effective at reducing the concentration of detectable gluten than each of the treatments alone. However, gluten proteins and/or polypeptides were observed in filtered, PEP-treated beers using sandwich ELISA methods, western blot, and bottom-up mass spectrometry. In addition, mass spectrometry results showed that the number of hydrolysed gluten peptides was almost unaffected by the filtration process. Gluten peptides that contained potentially immunopathogenic sequences were identified in the filtered PEP-containing beers by MS. Variability in gluten composition was observed between three replicate pilot-scale productions, suggesting that the gluten profile in beer could differ from batch to batch. As there is uncertainty in the detection and quantification of gluten in hydrolysed and fermented foods, characterisation of hydrolysed gluten by complementary analytical methodologies is recommended.