Novel column design for centrifugal countercurrent chromatography

设计了一种用于离心式逆流色谱柱的新型椭球柱。考察了3种不同的溶剂体系：正丁醇-冰醋酸-水(4：1：5，v/v)(BAW)、正己烷-乙酸乙酯-甲醇-0.1MHCl(1：1：1：1，v/v)(HEMH)和12.5%(w/w)聚乙二醇1000和12.5%(w/w)磷酸二氢钾水溶液(PEGDPP)。在BAW体系中，当柱角为0(柱轴与离心力平行)时，椭球柱的固定相保留率(SF)和峰分离度(Rs)均远高于90°(柱轴与离心力垂直)时的固定相保留率(Sf)和峰分离度(Rs)，其中在低流速下相洗脱时分离效果最好，当流速为0.07ml/min时，固定相保留率为2.02，峰分离度为27.8%。在HEMW系统分离DNP-氨基酸时，当流速为0.05ml/min时，31.6%的SF在DNP-DL-Glu和DNP--ALA峰之间以及DNP--ALA和DNP-L-ALA峰之间产生较高的Rs值，分别为2.16和1.81。在PEG-DPP体系中，溶菌酶和肌红蛋白的相对分子质量为1.08，流速为0.03ml/min，SF为38.9%。这些Rs值中的大多数都超过了以前报道的类似实验条件下从图8柱子中获得的值。 关键词：椭球柱，流体逆流色谱系统，离心逆流色谱，固定相保留，峰分辨，二肽，DNP-氨基酸，蛋白质

A novel ellipsoid column was designed for centrifugal counter-current chromatography. Performance of the ellipsoid column with a capacity of 3.4 mL was examined with three different solvent systems composed of 1-butanol-acetic acid-water (4:1:5, v/v) (BAW), hexane-ethyl acetate-methanol-0.1 M HCl (1:1:1:1, v/v) (HEMH), and 12.5% (w/w) PEG1000 and 12.5% (w/w) dibasic potassium phosphate in water (PEG-DPP) each with suitable test samples. In dipeptide separation with BAW system, both stationary phase retention (Sf) and peak resolution (Rs) of the ellipsoid column were much higher at 0 column angle (column axis parallel to the centrifugal force) than at 90 column angle (column axis perpendicular to the centrifugal force), where elution with the lower phase at a low flow rate produced the best separation yielding Rs at 2.02 with 27.8% Sf at a flow rate of 0.07 ml/min. In the DNP-amino acid separation with HEMW system, the best results were obtained at a flow rate of 0.05 ml/min with 31.6% Sf yielding high Rs values at 2.16 between DNP-DL-glu and DNP-β-ala peaks and 1.81 between DNP-β-ala and DNP-L-ala peaks. In protein separation with PEG-DPP system, lysozyme and myolobin were resolved at Rs of 1.08 at a flow rate of 0.03 ml/min with 38.9% Sf. Most of those Rs values exceed those obtained from the figure-8 column under similar experimental conditions previously reported. Keywords: ellipsoid column, hydrostatic counter-current chromatographic system, centrifugal counter-current chromatography, retention of the stationary phase, peak resolution, dipeptides, DNP-amino acids, proteins