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Open AccessArticle

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

1
Institute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Potsdam, Germany
2
IGV-Institut für Getreideverarbeitung GmbH, Arthur-Scheunert-Allee 40/41, 14558 Nuthetal, Germany
3
Department of Gastronomy, Faculty of Applied Science, University of Altınbaş, Büyükdere Cad. No 147, 34394 Esentepe-İstanbul, Turkey
*
Author to whom correspondence should be addressed.
Foods 2020, 9(10), 1448; https://doi.org/10.3390/foods9101448
Received: 28 August 2020 / Revised: 25 September 2020 / Accepted: 10 October 2020 / Published: 13 October 2020
(This article belongs to the Special Issue Analytical Methods for Allergen Control in Food Processing)
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. View Full-Text
Keywords: α-amylase/trypsin inhibitors; wheat cultivars; SDS-PAGE; peptides markers; relative quantification; mass spectrometry; LC-MRM-MS α-amylase/trypsin inhibitors; wheat cultivars; SDS-PAGE; peptides markers; relative quantification; mass spectrometry; LC-MRM-MS
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MDPI and ACS Style

Sagu, S.T.; Zimmermann, L.; Landgräber, E.; Homann, T.; Huschek, G.; Özpinar, H.; Schweigert, F.J.; Rawel, H.M. Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS. Foods 2020, 9, 1448. https://doi.org/10.3390/foods9101448

AMA Style

Sagu ST, Zimmermann L, Landgräber E, Homann T, Huschek G, Özpinar H, Schweigert FJ, Rawel HM. Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS. Foods. 2020; 9(10):1448. https://doi.org/10.3390/foods9101448

Chicago/Turabian Style

Sagu, Sorel T.; Zimmermann, Lynn; Landgräber, Eva; Homann, Thomas; Huschek, Gerd; Özpinar, Haydar; Schweigert, Florian J.; Rawel, Harshadrai M. 2020. "Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS" Foods 9, no. 10: 1448. https://doi.org/10.3390/foods9101448

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