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Selection of column and operating conditions for reversed phase high performance liquid chromatography of proteins in canadian wheat






Canadian Journal of Plant Science 65(2): 285-298

Selection of column and operating conditions for reversed phase high performance liquid chromatography of proteins in canadian wheat

Chromatographic conditions were optimized and 3 commercially available columns were evaluated for separation of alcohol-soluble storage proteins of 'Neepawa' wheat [Triticum aestivum] using reversed-phase high-performance liquid chromatography (RP-HPLC). Optimal separation was achieved using an extracting solution of 50% 1-propanol, 1% acetic acid and 4% dithiothreitol and an HPLC elution time of 105 min at a flow rate of 1.0 ml/min. HPLC columns evaluated (SynChropak RP-P, Ultrapore RPSC and Aquapore RP-300) varied in selectivity and resolution. The column providing the greatest versatility was Aquapore RP-300 available in cartridge form. Sodium dodecyl sulfate gradient-gel electrophoresis analysis of protein peaks resolved by RP-HPLC indicated that many of the eluted peaks contained morethan one protein species. Chromatographic protein patterns obtained for Neepawa wheat grown at different locations and in different years were qualitatively the same.


Accession: 006380990



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