Cereal Chem 62:332-339 | VIEW
Improved Procedures for Rapid Wheat Varietal Identification by Reversed-Phase High- Performance Liquid Chromatography of Gliadin.
J. A. Bietz and L. A. Cobb. Copyright 1985 by the American Association of Cereal Chemists, Inc.
Wheat varieties may be identified through reversed-phase high-performance liquid chromatography (RP- HPLC) of gliadins. RP-HPLC is fast, resproducible, and gives good resolution, but improvements are still possible by altering run time, flow rate, and temperature, and by using internal or external standards. Rapid gradients (10-15 min, 2.5-3.0 ml/min) gave less resolution than longer (about one hour) runs, but still differentiated most varieties, permitting more than 100 analyses per day per instrument. Reproducibility was improved by substituting H2O and CH3CN (+0.1% trifluoroacetic acid) for H2O/CH3CN/trifluoroacetic acid solvent blends used earlier. Constant temperature is essential for maximum reproducibility, and resolution is significantly better at elevated (70 C) temperature, possibly caused by better dissociation of proteins. Alkylphenones (C2-C10) may serve as standards to eliminate intra- and interlaboratory variability that arises from different experimental conditions. A hydrophobicity index is proposed to characterize proteins in relation to standards rather than by elution time. These modificatons for improving speed, resolution, and reproducibility of RP-HPLC gliadin analysis may permit automated wheat varietal identification by computer, free of subjective data evaluation.