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首页> 外文期刊>Journal of liquid chromatography and related technologies >Protein separation by cross-axis coil planet centrifuge with spiral column assemblies
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Protein separation by cross-axis coil planet centrifuge with spiral column assemblies

机译：交叉轴螺旋行星离心机与螺旋柱组件分离蛋白质

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摘要

A newly fabricated spiral column assembly was first applied to the countercurrent chromatographic separation of proteins using the cross-axis coil planet centrifuge (cross-axis CPC). The separation was performed by a set of stable proteins such as cytochrome C, myoglobin, and lysozyme with an aqueous-aqueous polymer phase solvent system composed of 12.5% (w/w) polyethylene glycol (PEG) 1000-12.5% (w/w) dibasic potassium phosphate. Three sets of left-handed single-layer spiral columns with different IDs (1.0, 1.5, and 2.0 mm) were employed to investigate the effect of column ID on partition efficiency at four different elution modes. The elution modes consisted of the combinations of the direction of revolution (PI = counterclockwise; PII = clockwise), the head-tail elution mode (H = head to tail; T = tail to head), and the inward-outward elution mode (I = inward; O = outward). Among these experiments, the best separation of proteins was attained using a 1.5 mm-ID column assembly with the PI-H-O elution mode of the lower phase mobile. The resolution between cytochrome C and myoglobin peaks was 0.9 and between myoglobin and lysozyme peaks was 1.0, while the retention of the stationary phase was 32.7%. In order to improve the partition efficiency, the four-layer spiral column assembly with 1.0 mm-ID column tubing was applied to the protein separation. The resolution between these peaks was improved to 1.1 and 1.2, respectively, while the stationary phase retention decreased to 20.9%. Further studies using a single-layer spiral coiled column also revealed the effective peak resolution of proteins, but with lower stationary phase retention. The overall results demonstrated the spiral column assembly was useful for the protein separation using the cross-axis CPC with the aqueous-aqueous polymer phase system.
机译：首先使用横轴线圈行星式离心机（横轴CPC）将一种新制造的螺旋柱组件应用于蛋白质的逆流色谱分离。分离是通过一组稳定的蛋白质（例如细胞色素C，肌红蛋白和溶菌酶）与由12.5％（w / w）1000-12.5％（w / w）的聚乙二醇（PEG）组成的水性聚合物相溶剂系统进行的）磷酸氢二钾。使用三组具有不同ID（1.0、1.5和2.0 mm）的左手单层螺旋柱，研究了四种不同洗脱模式下ID对分离效率的影响。洗脱模式由旋转方向（P I =逆时针; P II =顺时针），头尾洗脱模式（H =头尾相接）组成; T =从尾到头），以及从内到外的洗脱模式（I =向内; O =向外）。在这些实验中，使用1.5 mm-ID色谱柱组件和下相移动相的P I -H-O洗脱模式可实现蛋白质的最佳分离。细胞色素C和肌红蛋白峰之间的分辨率为0.9，而肌红蛋白和溶菌酶峰之间的分辨率为1.0，而固定相的保留率为32.7％。为了提高分配效率，将具有1.0 mm-ID内径管的四层螺旋柱组件应用于蛋白质分离。这些峰之间的分离度分别提高到1.1和1.2，而固定相保留率降低到20.9％。使用单层螺旋卷曲色谱柱的进一步研究还揭示了蛋白质的有效峰分离度，但固定相保留较低。总体结果表明，螺旋色谱柱组件可用于交叉轴CPC与水-水性聚合物相系统的蛋白质分离。

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