title: Separation and characterization of barley (Hordeum vulgare L.) hordeins by free zone capillary electrophoresis.
abstract: Extraction conditions, separation conditions, and capillary rinsing protocols were optimized for the separation of barley hordeins by free zone capillary electrophoresis. Stable hordein extracts were obtained with a single 5 min extraction after the albumins and globulins were removed. Hordeins had to be reduced for optimal resolution. Optimum separation conditions for hordein separations were 100 mM phosphate-glycine buffer containing 20% acetonitrile and 0.05% hydroxypropylmethylcellulose. The addition of zwitterionic sulfobetaine detergents containing hydrocarbon tails of eight and ten carbons slightly improved the resolution of the separations, but not enough to warrant their use on a routine basis. The migration positions of the hordein subclasses were determined by two- dimensional reversed-phase high-performance liquid chromatography x free zone capillary electrophoresis mapping. The hordein subclasses formed clusters similar to those of wheat gliadins. Separation-to-separation repeatability was good, with migration time relative standard deviations < 1% for a 15-run period. For routine discrimination of cultivars, a 2 min post-separation rinse with 500 mM acetic acid was necessary to prevent protein build-up on the capillary walls. An example of successfully differentiating barley cultivars using this technique is shown.
authors.affiliation: USDA-ARS, Grain Marketing and Production Research Center, and Kansas State University, Manhattan, USA. george@usgmrl.ksu.edu
authors.forename: G L
authors.initials: GL
authors.lastname: Lookhart
authors.forename: S R
authors.initials: SR
authors.lastname: Bean
authors.forename: B L
authors.initials: BL
authors.lastname: Jones
mesh_terms: D002851:Chromatography, High Pressure Liquid; D019075:Electrophoresis, Capillary / Q000379:methods*; D005983:Glutens; D001467:Hordeum / Q000737:chemistry*; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D013997:Time Factors
chemical_list: D010940:Plant Proteins; D005983:Glutens