Determination of Phosphoamino Acids by 6-Oxy-(N-succinimidyl acetate)-9-(2′-methoxycarbonyl)fluorescein and MEKC with LIF Detection |
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Authors: | Li-Wei Cao Wang Hong Hua-Shan Zhang |
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Institution: | 1. Department of Chemistry, Jinan University, Guangzhou, 510632, China 2. Department of Chemistry, Wuhan University, Wuhan, 430072, China
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Abstract: | The fluorescent reagent 6-oxy-(N-succinimidyl acetate)-9-(2′-methoxycarbonyl)fluorescein (SAMF), has been newly synthesized for use as a label for characterization of phosphoamino acids by micellar electrokinetic capillary electrophoresis (MEKC) with laser-induced fluorescence (LIF) detection. The conditions for derivatization and separation of the phosphoamino acids, including pH, concentration of electrolyte, and Brij-35 concentration were optimized in detail. Derivatization was performed at 35 °C for 10min in borate buffer (pH 8.0). The derivatives were separated to baseline by use of running buffer containing 50 mM borate and 20 mM Brij-35 at pH 9.3. Detection limits ranged from 5 × 10?11 to 1 × 10?9 mol L?1 (signal to noise ratio = 3). The method was used for characterization of the phosphoamino acids in a sample from hydrolysis of a novel protein kinase from tobacco cells. |
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