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首页> 外文期刊>Journal of Agricultural and Food Chemistry >Acetonitrile as a buffer additive for free zone capillary electrophoresis separation and characterization of maize (Zeamays L. ) and sorghum (Sorghum bicolor L. Moench) storage proteins.
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Acetonitrile as a buffer additive for free zone capillary electrophoresis separation and characterization of maize (Zeamays L. ) and sorghum (Sorghum bicolor L. Moench) storage proteins.

机译：乙腈作为缓冲添加剂用于玉米（Zeamays L.）和高粱（Sorghum bicolor L. Moench）存储蛋白的自由区毛细管电泳分离和表征。

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An improved method for separating and characterizing maize (Zea mays L.) and sorghum (Sorghum bicolor L. Moench) storage proteins by free zone capillary electrophoresis (FZCE) was developed. Previous electrophoretic methods for analyzing these proteins required high concentrations of urea to maintain protein solubility during separation. To overcome disadvantages of urea, we developed a FZCE method that mimicked reversed-phase high-performance liquid chromatography (RP-HPLC) in that it used high levels of acetonitrile (ACN) at low pH. The optimized FZCE buffer system consisted of 80 mM phosphate-glycine buffer, nominal pH 2.5, containing 60% ACN and a cellulose derivative to dynamically coat capillary walls. Resolution was similar to or higher than that previously achieved by FZCE buffers utilizing 8 M urea as a buffer additive. ACN concentrations of at least 50% were necessary to achieve acceptable separations; this ACN concentration is approximately that necessary to extract these storage proteins. ACN was equally effective as traditional ethanol solvents and 8 M urea for solubilizing maize and sorghum proteins. The ACN-based FZCE buffer system gave high repeatability (<0.3% relative standard deviation, measured over 15 consecutive injections) for migration time. Subclasses of maize and sorghum storage proteins were identified, and genotypes of each cereal were successfully differentiated using ACN-containing buffers. This FZCE method may be applicable for the analysis of other hydrophobic proteins without the use of urea.
机译：开发了一种通过自由区毛细管电泳（FZCE）分离和鉴定玉米（Zea mays L.）和高粱（Sorghum bicolor L. Moench）贮藏蛋白的方法。用于分析这些蛋白质的以前的电泳方法需要高浓度的尿素以在分离过程中保持蛋白质溶解度。为了克服尿素的缺点，我们开发了一种FZCE方法，该方法模仿了反相高效液相色谱（RP-HPLC），因为它在低pH值下使用了高含量的乙腈（ACN）。优化的FZCE缓冲液系统由标称pH 2.5的80 mM磷酸盐-甘氨酸缓冲液组成，含有60％的ACN和纤维素衍生物以动态覆盖毛细管壁。分辨率与之前使用8 M尿素作为缓冲添加剂的FZCE缓冲液所达到的分辨率相似或更高。为了达到可接受的分离水平，必须使ACN浓度至少达到50％;该ACN浓度大约是提取这些存储蛋白所需的浓度。 ACN与传统的乙醇溶剂和8 M尿素在溶解玉米和高粱蛋白质方面同样有效。基于ACN的FZCE缓冲系统在迁移时间上具有很高的重复性（相对标准偏差<0.3％，在15次连续进样中测得）。确定了玉米和高粱贮藏蛋白的亚类，并使用含ACN的缓冲液成功区分了每种谷物的基因型。这种FZCE方法可适用于不使用尿素的其他疏水蛋白的分析。

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