Author
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Huebner, Floyd |
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Submitted to: American Association of Cereal Chemists Meetings
Publication Type: Abstract Only Publication Acceptance Date: 10/12/1997 Publication Date: N/A Citation: N/A Interpretive Summary: Technical Abstract: Some of the best early separations of wheat gliadins were achieved by ion-exchange column chromatography on a strong cation exchanger, sulfoethyl cellulose, eluting proteins with a salt gradient. Ten to 13 fractions were typically isolated in sufficient quantities for further analyses, but the method was laborious and time-consuming. The advent of high-performance liquid chromatography (HPLC) introduced greatly-improved fractionation procedures based on hydrophobicity and size, but few satisfactory ion-exchange HPLC columns were available. A new ion-exchange HPLC column similar in nature to those cellulosic columns originally used to fractionate gliadin has now become available and has been tested for its ability to fractionate this protein mixture. The column is a Vydac sulfonic acid S-type cation (7.5 x 50 mm). This column gives separations almost as good as those of reversed-phase HPLC: more than 30 components resolve. A salt gradient was used to elute the gliadins from 0 to 0.25M NaCl in a solvent consisting of 30 to 35% acetonitrile, and 0.05% TFA adjusted to pH of 2.6. A run time of 45 min was used. Individual IE-HPLC peaks were collected for subsequent analysis and characterization by RP-HPLC and gel electrophoresis, revealing which gliadins elute in each peak. Analyses of glutenin subunits (not yet fully explored) were also achieved. Wheat varieties could be readily differentiated by the resulting separations and relationships of protein composition to quality were apparent. Proteins from other cereals were also successfully separated. It appears that this IE-HPLC procedure may provide another useful and complementary method for isolation and characterization of cereal proteins. |
