0000000000280401
AUTHOR
J.n. Kinkel
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…
Evaluation of advanced silica packings for the separation of biopolymers by high-performance liquid chromatography
Abstract The reversed-phase chromatography of proteins by gradient elution with acidic, low-ionic-strength aqueous—organic eluents is often associated with losses of the biological activity of the protein. In this study, the enzymatic activities of catalase, horseradish peroxidase and pepsin were examined under static and dynamic column conditions on non-porous, monodisperse 1.5-μm reversed-phase silicas with various n -alkyl ligands. Catalase readily lost its enzymatic activity under the influence of the acidic aqueous—organic eluents in the absence of the reversed-phase packing, whereas peroxidase was partially deactivated as a result of combined mobile phase and stationary phase effects …
Packing technology, column bed structure and chromatographic performance of 1-2-μm non-porous silicas in high-performance liquid chromatography
This work is aimed at further elucidating the aggregation behaviour of micron- and submicron-size non-porous silicas and the column performance of 1–2-μm C18 silicas in reversed-phase high-performance liquid chromatography of low-molecular weight compounds. It is demonstrated that highly ordered, dense, porous aggregates of such silica beads were obtained by gravity settling and centrifugation. The slurry techniques applied at constant flow-rate and a pressure up to 50 MPa provided less-ordered aggregates, but generated an acceptable performance of columns when 1–2-μm C18 silica beads were employed. To operate columns of 53 mm × 4.6 mm I.D., the maximum flow-rate needs to be ca. 2.5 ml/min …
Nonporous Silica Microspheres in the Micron and Submicron Size Range
Uniform nonporous silica microspheres in the micrometer and submicrometer size range are formed by a process which involves reacting alkoxysilanes with water in a short chain alcohol using dissolved ammonia as a morphological catalyst.
Chapter 8 Native and Bonded Silicas in Aqueous Sec
Publisher Summary This chapter deals with the properties of packings and columns in correlation to chromatographic separations in size exclusion chromatography (SEC) and provides guidelines for optimum support design. The chapter provides a survey of the current state-of-the-art with respect to the surface modification of silica for SEC packings in aqueous eluents. The properties of commercial packings and columns are listed and compared. An advice is also discussed on the operation and maintenance of columns with a view to achieving high resolution and long life-time. In SEC, solute elution is based on equilibrium, entropy-controlled size exclusion process. According to their size, the so…
Polymer support synthesis
Non-porous silica gel microbeads of diameter 1.5 microns have been investigated as supports for oligonucleotide synthesis. In the preparation of oligothymidylates of chain length up to 150 bases, with 5'-di-p-anisylphenylmethyl-3'-phosphoramidite as an intermediate, the average yields per chain elongation were up to 99%. Lower overall yields were observed in the case of a support which developed a strong tendency towards aggregation after the build up of an oligonucleotide coating.
Evaluation of advanced silica packings for the separation of biopolymers by high-perforamnce liquid chromatography
Non-porous monodisperse 1.5-μm silicas were allowed to react with (A) and (B) N-acetylaminopropyltriethoxysilane to generate bonded phases useful in high-performance hydrophobic-interaction chromatography (HIC). Differences in the selectivity were observed between he amide and the ether phase. Peak capacities between 10 and 30 were achieved for several proteins with the amide and ether phase packed into columns of 36 × 8 mm I.D. and elution of the proteins under chromatographic conditions in which the gradient volume, VG, was held constant by varying the gradient time between 20 and 2.5 min and the flow-rate between 0.5 and 4.0 ml/min. The S values derived from the dependences of log k′ on …