Evaluation of advanced silica packings for the separation of biopolymers by high-performance liquid chromatography

Following previous studies of the use of non-porous monodisperse 1.5-microns n-octyl- and n-octadecyl-bonded silicas in gradient elution of proteins, this work was aimed at elucidating further the properties of this novel column material for peptide and protein separations in comparison with wide-pore silicas. First, it is demonstrated that with short columns (e.g., 35 X 8 mm I.D.) packed with these non-porous reversed-phase materials, mixtures of small peptides and mixtures of proteins can be very efficiently resolved. When the chain length of the bonded ligand was varied, the retention of a test set of proteins in gradient elution followed the ligand sequence C18 greater than C8 approxima…

Packings and stationary phases for biopolymer separations by HPLC

Packings and stationary phases applied to high resolution separations of proteins, enzymes, and nucleic acids must satisfy a series of distinct criteria that are different from those usually required by HPLC of low molecular weight non-biologically active analytes. These requirements have been met through substantial improvements in classical gel media together with novel developments in silica supports, and have led to a family of products with tailor-made and reproducible properties. Supports consisting of cross-linked organic gels, and inorganic materials (mostly silicas) are now available with graduated particle sizes, pore sizes, porosities and surface areas as well as non-porous beads…

Non-porous microparticulate supports in high-performance liquid chromatography (HPLC) of biopolymers — concepts, realization and prospects

BONDED SILICA PHASES FOR THE SEPARATION OF BIOPOLYMERS BY MEANS OF COLUMN LIQUID CHROMATOGRAPHY

Evaluation of advanced silica packings for the separation of biopolymers by high-performance liquid chromatography

Abstract The linear solvent strength model of Snyder was applied to describe fast protein separations on 2.1-μm non-porous, silica-based strong anion exchangers. It was demonstrated on short columns packed with these anion exchangers that (i) a substantially higher resolution of proteins and nucleotides was obtained at gradient times of less than 5 min than on porous anion exchangers; (ii) the low external surface area of the non-porous anion exchanger is not a critical parameter in analytical separations and (iii) μg-amounts of enzymes of high purity and full biological activity were isolated.

Optimisation of fast protein separations on non-porous silica-based strong anion exchangers

The adsorbed coating technology using various vinylpyrrolidone-vinylimidazole copolymer compositions was carried out on 1.7 μm non-porous monodisperse silica. It was shown that the retention properties and the loading capacity for bovine serum albumin (BSA) increases with the amount of vinylimidazole in the copolymer composition. The retention behavior of various proteins as a function of the salt composition in the eluent has been applied to find the optimal conditions for the synthesis of the anion-exchange stationary phase. The suitability of these supports for the fast separation of biological molecules is demonstrated. The best resolution and the highest speed in protein analysis was o…