Partition efficiency of the high-pitch locular multilayer coil was evaluated in countercurrent chromatographic (CCC) separation of proteins with an aqueous-aqueous polymer phase system using the small-scale cross-axis coil planet centrifuge (X-axis CPC) fabricated in our laboratory. The separation column was specially made by high-pitch (ca 5 cm) winding of 1.0 mm I.D., 2.0 mm O.D. locular tubing compressed at 2 cm intervals with a total capacity of 29.5 mL. The protein separation was performed using a set of stable proteins including cytochrome C, myoglobin, and lysozyme with the 12.5% (w/w) polyethylene glycol (PEG) 1000 and 12.5% (w/w) dibasic potassium phosphate system (pH 9.2) under 1000 rpm of column revolution. This high-pitch locular tubing yielded substantially higher stationary phase retention than the normal locular tubing for both lower and upper mobile phases. In order to demonstrate the capability of the high-pitch locular tubing, the purification of collagenase from the crude commercial sample was carried out using an aqueous-aqueous polymer phase system. Using the 16.0% (w/w) PEG 1000–6.3% (w/w) dibasic potassium phosphate–6.3% (w/w) monobasic potassium phosphate system (pH 6.6), collagenase I, II, IV, V, and X derived from Clostridium histolyticum were separated from other proteins and colored small molecular weight compounds present in the crude commercial sample. Collagenase from C. histolyticum well retained its enzymatic activity in the purified fractions. The overall results demonstrated that the high-pitch locular multilayer coil is effectively used for the CCC purification of bioactive compounds without loss of their enzymatic activities.

中文翻译：

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使用小规模横轴线圈行星离心机对蛋白质进行逆流色谱分离的高螺距局部多层线圈的分离效率以及在使用水相水酶系统纯化各种胶原蛋白中的应用

使用我们实验室制造的小型交叉轴螺旋行星离心机（X 轴 CPC），在具有水-水聚合物相系统的蛋白质逆流色谱 (CCC) 分离中评估高螺距室状多层线圈的分配效率. 分离柱是由内径为 1.0 毫米、外径为 2.0 毫米的腔管以 2 厘米间隔压缩的高节距（约 5 厘米）缠绕而成，总容量为 29.5 毫升。使用一组稳定的蛋白质进行蛋白质分离，包括细胞色素 C、肌红蛋白和溶菌酶以及 12.5% (w/w) 聚乙二醇 (PEG) 1000 和 12.5% (w/w) 磷酸氢钾系统 (pH 9.2)低于 1000 rpm 的色谱柱旋转。对于较低和较高的流动相，这种高螺距室管产生的固定相保留率明显高于普通室管。为了证明高螺距室管的能力，使用水-水聚合物相系统从粗商业样品中纯化胶原酶。使用 16.0% (w/w) PEG 1000–6.3% (w/w) 磷酸氢二钾–6.3% (w/w) 磷酸二氢钾系统 (pH 6.6)、胶原酶 I、II、IV、V 和 X来源于溶组织梭菌的细菌与其他蛋白质和存在于粗商业样品中的有色小分子量化合物分离。来自溶组织梭菌的胶原酶在纯化的级分中很好地保留了其酶活性。