The use of reversed-phase high-performance liquid chromatography with radial compression for the analysis of peptide and protein mixtures.

This report describes the separation of peptide and protein mixtures on a C18- microparticulate support which was packed in a polyethylene cartridge and subjected to radial compression of ca. 2600 p.s.i. The purpose of the radial compression was to minimise inhomogeneities in the column packing, in particular in the region of the column wall and end fittings. The effectiveness of this new chromatographic system was demonstrated by the efficient separation of the following mixtures: the C-apolipoproteins isolated from human very low density lipoproteins; the polymorphs of apolipoprotein A-I; the tryptic fragments of apolipoprotein C-II; the complex mixture generated by partial proteolysis of apolipoprotein B; the tryptic fragments of 3H- and 14C-labelled beta-chain of murine IA alloantigen and the tryptic fragments of carboxymethylated lambda chain isolated from human immunoglobulin G. The separated peaks were identified by amino acid analysis, radioactivity counting and in the last example by amino acid sequence determination. The mobile phase consisted of 1% aqueous solution of triethylammonium phosphate, pH 3.2 with acetonitrile or isopropanol as the organic modifier.