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Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS

The α-amylase/trypsin inhibitors (ATIs) are discussed as being responsible for non-celiac wheat sensitivity (NCWS), besides being known as allergenic components for baker’s asthma. Different approaches for characterization and quantification including proteomics-based methods for wheat ATIs have bee...

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Detalles Bibliográficos
Autores principales: Sagu, Sorel Tchewonpi, Zimmermann, Lynn, Landgräber, Eva, Homann, Thomas, Huschek, Gerd, Özpinar, Haydar, Schweigert, Florian J., Rawel, Harshadrai M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7600490/
https://www.ncbi.nlm.nih.gov/pubmed/33066015
http://dx.doi.org/10.3390/foods9101448
Descripción
Sumario:The α-amylase/trypsin inhibitors (ATIs) are discussed as being responsible for non-celiac wheat sensitivity (NCWS), besides being known as allergenic components for baker’s asthma. Different approaches for characterization and quantification including proteomics-based methods for wheat ATIs have been documented. In these studies generally the major ATIs have been addressed. The challenge of current study was then to develop a more comprehensive workflow encompassing all reviewed wheat-ATI entries in UniProt database. To substantially test proof of concept, 46 German and Turkish wheat samples were used. Two extractions systems based on chloroform/methanol mixture (CM) and under buffered denaturing conditions were evaluated. Three aspects were optimized, tryptic digestion, chromatographic separation, and targeted tandem mass spectrometric analysis (HPLC-MS/MS). Preliminary characterization with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) documented the purity of the extracted ATIs with CM mixture and the amylase (60–80%)/trypsin (10–20%) inhibition demonstrated the bifunctional activity of ATIs. Thirteen (individual/common) biomarkers were established. Major ATIs (7–34%) were differently represented in samples. Finally, to our knowledge, the proposed HPLC-MS/MS method allowed for the first time so far the analysis of all 14 reviewed wheat ATI entries reported.