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 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 amylaseThe 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.…
Verfasserangaben: | Sorel Sagu TchewonpiORCiD, Lynn Zimmermann, Eva Landgräber, Thomas HomannORCiD, Gerd HuschekORCiDGND, Haydar Özpinar, Florian J. SchweigertORCiDGND, Harshadrai Manilal RawelORCiDGND |
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URN: | urn:nbn:de:kobv:517-opus4-486118 |
DOI: | https://doi.org/10.25932/publishup-48611 |
ISSN: | 1866-8372 |
Titel des übergeordneten Werks (Deutsch): | Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe |
Schriftenreihe (Bandnummer): | Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe (1028) |
Publikationstyp: | Postprint |
Sprache: | Englisch |
Datum der Erstveröffentlichung: | 10.12.2020 |
Erscheinungsjahr: | 2020 |
Veröffentlichende Institution: | Universität Potsdam |
Datum der Freischaltung: | 10.12.2020 |
Freies Schlagwort / Tag: | LC-MRM-MS; SDS-PAGE; mass spectrometry; peptides markers; relative quantification; wheat cultivars; α-amylase/trypsin inhibitors |
Ausgabe: | 1028 |
Seitenanzahl: | 27 |
Quelle: | Foods 9 (2020) 10 Art. 1448 DOI: 10.3390/foods9101448 |
Organisationseinheiten: | Mathematisch-Naturwissenschaftliche Fakultät / Institut für Ernährungswissenschaft |
DDC-Klassifikation: | 6 Technik, Medizin, angewandte Wissenschaften / 63 Landwirtschaft / 630 Landwirtschaft und verwandte Bereiche |
6 Technik, Medizin, angewandte Wissenschaften / 64 Hauswirtschaft und Familie / 640 Hauswirtschaft und Familie | |
Peer Review: | Referiert |
Publikationsweg: | Open Access / Green Open-Access |
Externe Anmerkung: | Bibliographieeintrag der Originalveröffentlichung/Quelle |