Efficiency of Poroshell type chromatogrphic columns for wheat gliadin separation using RP-HPLC
Jacek Waga
j.waga@ihar.edu.plInstytut Hodowli i Aklimatyzacji Roślin — PIB (Poland)
Jerzy Zientarski
Instytut Hodowli i Aklimatyzacji Roślin — PIB (Poland)
Maciej Szaleniec
Instytut Katalizy i Fizykochemii Powierzchni im. Jerzego Habera PAN (Poland)
Andrzej Skoczowski
Instytut Fizjologii Roślin im. Franciszka Górskiego PAN (Poland)
Abstract
RP-HPLC is an efficient tool for wheat storage proteins separation. Its extensive utilization in plant breeding is limited, however, by heavy expenses of apparatus, equipment and reagents. Recently a new type of chromatographic columns was introduced where stationary phase is based on the superficially porous silca (so called Fused-Core Particles). The main advantage of the Fused-Core stationary phase is a high increase of separation power (theoretical plates) which enables shortening of the separation time (up to few minutes) and consequently the reduction of the analysis costs. The aim of this study was a comparative evaluation of the accuracy and efficacy of the Fused-Core C18 column with a classic C18 column in the separation of wheat gliadins — one of the most complex and polymorphic plant proteins. Closely related winter wheat hybrid lines, differentiated in respect of gliadin controlling alleles, were analysed. Gentypes containing gliadin null alleles and corresponding control lines as well as parental forms and their progenies were compared using Fused-Core Poroshell 300SB-C18 and traditional Zorbax 300SB-C18 columns. The obtained results proved that RP-HPLC separation on the Poroshell column are indeed very fast and economical as the high quality chromatograms were obtained after 8 minutes of separation. However, their resolution was considerably lower as compared to the traditional column. In conclusion, the rapid PR-HPLC appears to be very well suited for mass selection of breeding materials, where large number of genotypes must be evaluated in a very short time period. However in a genetic research, where identification of discrete details of the chromatograms could be important for genotype classification, traditional columns seem to be more useful.
Supporting Agencies
Keywords:
RP-HPLC, Poroshell, Zorbax C18, wheat gliadin proteinsReferences
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Authors
Jerzy ZientarskiInstytut Hodowli i Aklimatyzacji Roślin — PIB Poland
Authors
Maciej SzaleniecInstytut Katalizy i Fizykochemii Powierzchni im. Jerzego Habera PAN Poland
Authors
Andrzej SkoczowskiInstytut Fizjologii Roślin im. Franciszka Górskiego PAN Poland
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