Spiral Countercurrent Chromatography Studies Using the Spiral Disk Assembly

Abstract

The first rotor proposed for high speed countercurrent chromatography (HSCCC), the spiral disk assembly, consisting of a stack of 8 high density polyethylene plates with spiral flow channels, has been studied with various solvents and peptide separations to understand its application to polar compounds and solvent systems. Mostly polar solvent systems comprised of the heavy alcohols, were studied because of their suitability for peptide and larger molecule separation. Peptide mixtures were separated to correlate partition coefficients ranging from 0.3 to 2.8 with elution in spiral CCC. With some peptides, scale-up purification experiments were performed with sample loadings up to 85 mg in the 153 mL volume rotor. These studies characterized polar solvent systems with high stationary phase retentions that can be used for separation and purification of compounds with suitable partition coefficients. The solvent system, sec-butanol-1% aq.trifluoroacetic acid with a 73.3% stationary phase retention proved to be useful for most separations. This is a very useful method to substitute for the use of acetonitrile in preparative chromatography.