Continuous beds for microchromatography: reversed-phase chromatography.

Simple and cost-effective methods for the preparation of microcolumns (i.d. 0.025-0.32 mm) for reversed-phase chromatography are described. The procedure includes (1) synthesis in the column tube of a continuous bed matrix from a monomer solution (piperazine diacrylamide, methacrylamide) containing allyl glycidyl ether and 2-hydroxyethyl methacrylate and (2) linking of C18 ligands by reacting 1,2-epoxyoctadecane with the epoxy and hydroxy groups in the matrix. The derivatization can be accomplished within 20 min. The columns prepared in this way showed high performance in the separation of proteins and peptides and permitted short analysis times (100 s).