Next Article in Journal
Previous Article in Journal
Previous Article in Special Issue
Nutrients 2014, 6(4), 1578-1597; doi:10.3390/nu6041578
Article

Analysis of Wheat Prolamins, the Causative Agents of Celiac Sprue, Using Reversed Phase High Performance Liquid Chromatography (RP-HPLC) and Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS)

1,†,‡
, 1,†
, 1
, 2
, 1,3,4,*  and 1,*
Received: 27 December 2013; in revised form: 26 March 2014 / Accepted: 28 March 2014 / Published: 15 April 2014
(This article belongs to the Special Issue Dietary Protein and Human Health)
View Full-Text   |   Download PDF [2453 KB, uploaded 15 April 2014]   |   Browse Figures
Abstract: Wheat prolamins, commonly known as “gluten”, are a complex mixture of 71–78 proteins, which constitute ~80% of the proteins in the wheat grains and supply 50% of the global dietary protein demand. Prolamins are also responsible for numerous gluten-induced disorders and determine the unique visco-elastic properties of the wheat dough. These properties necessitate the reliable determination of the prolamin composition in wheat grains and their derived products. Therefore, this study examined the impact of HPLC conditions, including column type, column temperature, flow rate, and the gradient of polar and non-polar solvents in the mobile phase, to improve the analytical resolution of prolamins. The following conditions were found optimal for analyses: column temperature 60 °C, flow rate 1.0 mL/min and an elution gradient of 20%–60% of 0.1% trifluoroacetic acid + acetonitrile in 60 min. For further improvement of resolution, gliadin and glutenin extracts were analyzed using MALDI-TOF-MS in combination with HPLC fractionation. Two semi-quantitative methods, densitometry of stained polyacrylamide gels and HPLC, were used to determine relative prolamin quantities and the correspondence between the methods was established. The combinatorial gluten analyses approach developed during the present study was used to analyze prolamin profiles of wheat transformants expressing DEMETER silencing artificial microRNA, and the results are discussed.
Keywords: prolamins; glutenin; gliadin; celiac disease; wheat prolamins; glutenin; gliadin; celiac disease; wheat
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Export to BibTeX |
EndNote


MDPI and ACS Style

Mejías, J.H.; Lu, X.; Osorio, C.; Ullman, J.L.; von Wettstein, D.; Rustgi, S. Analysis of Wheat Prolamins, the Causative Agents of Celiac Sprue, Using Reversed Phase High Performance Liquid Chromatography (RP-HPLC) and Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS). Nutrients 2014, 6, 1578-1597.

AMA Style

Mejías JH, Lu X, Osorio C, Ullman JL, von Wettstein D, Rustgi S. Analysis of Wheat Prolamins, the Causative Agents of Celiac Sprue, Using Reversed Phase High Performance Liquid Chromatography (RP-HPLC) and Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS). Nutrients. 2014; 6(4):1578-1597.

Chicago/Turabian Style

Mejías, Jaime H.; Lu, Xiaoqiao; Osorio, Claudia; Ullman, Jeffrey L.; von Wettstein, Diter; Rustgi, Sachin. 2014. "Analysis of Wheat Prolamins, the Causative Agents of Celiac Sprue, Using Reversed Phase High Performance Liquid Chromatography (RP-HPLC) and Matrix-Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS)." Nutrients 6, no. 4: 1578-1597.



Nutrients EISSN 2072-6643 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert