COLUMN CONFIGURATION FOR PROTEIN SEPARATION--TOROIDAL CENTRIFUGE CHROMATOGRAPHY

点击翻译按钮获取中文摘要

The toroidal coil centrifuge was used for protein separation with polymer phase systems by countercurrent chromatography. Two series of basic studies were conducted: in the first series of experiments three different types of separation column were constructed to evaluate the performance of each column for partition of proteins using a polymer phase system composed of polyethylene glycol 1000 and dibasic potassium phosphate, each at 12.5% (w/w). In each experiment the column was first entirely filled with the upper PEG-rich upper stationary phase followed by injection of the sample solution. Then, the phosphate-rich lower mobile phase was eluted through the column at 0.2ml/min. The fractions (each 0.4 ml) were analyzed by spectrophotometer at 280 nm to obtain the elution curve. The partition efficiency was determined from the chromatogram using a conventional equation. The result indicated that the toroidal coil yielded considerably higher efficiency than the twisted column, while the efficiency of the radial coil arrangement was lowest In the second series of our studies, both toroidal and twisted coils were tested to detect the effect of the Coriolis force by switching the inlet and outlet of the column and using both upper and the lower phases as the mobile phase. The results showed that the partition efficiency was improved when the effective side of the coil was placed near parallel to the direction of the Coriolis force. This tendency was more pronounced by reducing the core diameter and the toroidal coil prepared from 3 mm core, if eluted in the suitable direction, yielded the highest partition efficiency. The effect of the Coriolis force on the countercurrent chromatographic separation has not been reported previously. This Coriolis effect, however, was not observed on the twisted coil. Also we have found that a similar experiment using the organic/aqueous solvent system in the toroidal coil showed no Coriolis effect on partition efficiency.