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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| Format: | Article |
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MDPI AG
2020-10-01
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| Series: | Foods |
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| Online Access: | https://www.mdpi.com/2304-8158/9/10/1448 |
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| author | Sorel Tchewonpi Sagu Lynn Zimmermann Eva Landgräber Thomas Homann Gerd Huschek Haydar Özpinar Florian J. Schweigert Harshadrai M. Rawel |
| author_facet | Sorel Tchewonpi Sagu Lynn Zimmermann Eva Landgräber Thomas Homann Gerd Huschek Haydar Özpinar Florian J. Schweigert Harshadrai M. Rawel |
| author_sort | Sorel Tchewonpi Sagu |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-03-10T15:41:30Z |
| format | Article |
| id | doaj.art-7d3d833375344ee2862448736410cb40 |
| institution | Directory Open Access Journal |
| issn | 2304-8158 |
| language | English |
| last_indexed | 2024-03-10T15:41:30Z |
| publishDate | 2020-10-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Foods |
| spelling | doaj.art-7d3d833375344ee2862448736410cb402023-11-20T16:49:29ZengMDPI AGFoods2304-81582020-10-01910144810.3390/foods9101448Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MSSorel Tchewonpi Sagu0Lynn Zimmermann1Eva Landgräber2Thomas Homann3Gerd Huschek4Haydar Özpinar5Florian J. Schweigert6Harshadrai M. Rawel7Institute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Potsdam, GermanyInstitute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Potsdam, GermanyInstitute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Potsdam, GermanyInstitute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Potsdam, GermanyIGV-Institut für Getreideverarbeitung GmbH, Arthur-Scheunert-Allee 40/41, 14558 Nuthetal, GermanyDepartment of Gastronomy, Faculty of Applied Science, University of Altınbaş, Büyükdere Cad. No 147, 34394 Esentepe-İstanbul, TurkeyInstitute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Potsdam, GermanyInstitute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Potsdam, GermanyThe 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.https://www.mdpi.com/2304-8158/9/10/1448α-amylase/trypsin inhibitorswheat cultivarsSDS-PAGEpeptides markersrelative quantificationmass spectrometry |
| spellingShingle | Sorel Tchewonpi Sagu Lynn Zimmermann Eva Landgräber Thomas Homann Gerd Huschek Haydar Özpinar Florian J. Schweigert Harshadrai M. Rawel Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS Foods α-amylase/trypsin inhibitors wheat cultivars SDS-PAGE peptides markers relative quantification mass spectrometry |
| title | Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS |
| title_full | Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS |
| title_fullStr | Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS |
| title_full_unstemmed | Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS |
| title_short | Comprehensive Characterization and Relative Quantification of α-Amylase/Trypsin Inhibitors from Wheat Cultivars by Targeted HPLC-MS/MS |
| title_sort | comprehensive characterization and relative quantification of α amylase trypsin inhibitors from wheat cultivars by targeted hplc ms ms |
| topic | α-amylase/trypsin inhibitors wheat cultivars SDS-PAGE peptides markers relative quantification mass spectrometry |
| url | https://www.mdpi.com/2304-8158/9/10/1448 |
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