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PubMed2000

Simultaneous separation and quantitation of the major bovine whey proteins including proteose peptone and caseinomacropeptide by reversed-phase high-performance liquid chromatography on polystyrene-divinylbenzene.

A precise, sensitive and reliable RP-HPLC method was developed to enable not only unequivocal determination of alpha-lactalbumin and beta-lactoglobulin in bovine whey samples, but also simultaneous measurement of proteose peptone, caseinomacropeptide, bovine serum albumin and immunoglobulin G. The optimised method on the Resource RPC column allowed separation of the proteins in 30 min and could be applied to the analysis of soluble proteins in a variety of commercial and laboratory whey products. Furthermore, some qualitative information on protein heterogeneity and quality could be derived from the RP-HPLC analyses with additional data available from on-line electrospray mass spectrometry. Within- and between-day repeatability over a wide range of concentrations was excellent (RSD< or =5%) for all proteins except immunoglobulin G and bovine serum albumin where RSD was 7-10%. Analysis of grouped data from whey protein concentrate and whey protein isolate samples gave a limit of detection of < or =0.3% powder mass and a limit of quantitation of < or =1.0% powder mass for all proteins except immunoglobulin G. Limits of detection and quantitation were 0.6% and 2.0%, respectively, for this protein. Quantitative data obtained by the RP-HPLC method compared very favourably with data obtained by alternative methods of whey protein analysis.

Amino Acids↗
PubMed1988

Synthesis of a non-porous, polystyrene-based strong anion-exchange packing material and its application to fast high-performance liquid chromatography of proteins.

Adsorbed coating technology has been used to produce a strong anion-exchange stationary phase on 3-micron non-porous poly(styrene-divinylbenzene) particles. In order to take full advantage of the excellent kinetic properties of the resultant packing material, small columns of 5 mm X 6 mm I.D. were used. These columns were pressure- and pH-stable and allowed protein separations to be made in less than 1 min at ambient temperature.

Chemical Phenomena↗
PubMed2002

Detection and quantitation of lactoferrin in bovine whey samples by reversed-phase high-performance liquid chromatography on polystyrene-divinylbenzene.

An existing RP-HPLC method for the measurement of the major bovine whey proteins in bovine whey samples and powders has been extended to include the analysis of the minor bovine whey protein lactoferrin. Lactoferrin could be detected and quantitated at levels down to 0.2 microg and linear calibration was observed between 0.2 and 30 microg. Reliable quantitation of lactoferrin in whey samples could be achieved provided the bovine serum albumin to lactoferrin ratio did not exceed 10:1. Quantitative data obtained by the RP-HPLC method compared favourably with data obtained by Mono S analytical chromatography.

Animals↗
PubMed1991

Coated hydrophilic polystyrene-based packing materials.

A very hydrophilic high-performance liquid chromatographic base support was created from microparticulate, macroporous poly(styrene-divinylbenzene) beads. An organic monomer containing cross-linking functionalities was coated on the poly(styrene-divinylbenzene), followed by a catalyzed cross-linking reaction. The coatings formed contain only stable chemical bonds (e.g., C-C, C-O-C), and easily-derivatized hydroxyl moieties. This coated base support was evaluated for hydrophilicity, chemical stability, solvent compatibility, rigidity, and irreversible adsorption. Derivatives of the coated base support were made and applied in various modes of chromatography.

Cations↗
PubMed2002

High-performance liquid chromatographic separation of carbohydrates on a stationary phase prepared from polystyrene-based resin and novel amines.

New anion-exchange stationary phases On (n = 1, 2 and 3) with a dimethylamino terminal functional group, where n is the number of oxyethylene units [-(CH2CH2O)n-], were prepared by the reaction of chloromethylated porous styrene-divinylbenzene copolymer beads and amines [(CH3)2N-(CH2CH2O)nCH2CH2-N(CH3)2]. HPLC separations of monosaccharides (sorbitol, fucose, glucosamine, mannose, glucose, galactose, fructose, allose and altrose) and disaccharides (trehalose, lactose, cellobiose and maltose) were performed successfully on these stationary phases. The ether group of the stationary phases On was found to affect the separation of carbohydrates.

Amines↗
PubMed2002

Solid-phase extraction sorbent consisting of alkyltrimethylammonium surfactants immobilized onto strong cation-exchange polystyrene resin.

Presented is a solid-phase extraction sorbent material composed of cationic alkyltrimethylammonium surfactants attached to a strong cation-exchange resin via ion-exchange. The original hydrophilic cation-exchange resin is made hydrophobic by covering the surface with alkyl chains from the hydrophobic portion of the surfactant. The sorbent material now has a better ability to extract hydrophobic molecules from aqueous samples. The entire stationary phase (alkyltrimethylammonium surfactant) is removed along with the analyte during the elution step. The elution step requires a mild elution solvent consisting of 0.25 M Mg2+ in a 50% 2-propanol solution. The main advantage of using a removable stationary phase is that traditionally utilized toxic elution solvents such as methylene chloride, which are necessary to efficiently release strongly hydrophobic species from SPE stationary phases, may now be avoided. Also, the final extract is directly compatible with reversed-phase liquid chromatography. The performance of this procedure is presented using pyrene as a test molecule.

Cation Exchange Resins↗
PubMed2003

Evaluation of the two-dimensional reversed-phase-reversed-phase separations of low-molecular mass polystyrenes.

The resolving power of four reversed-phase-reversed-phase two-dimensional (2D) chromatographic systems was evaluated using information theory (IT) and a geometric approach to factor analysis. The first separation dimension employed a C18 column, while the second separation dimension employed a carbon clad zirconia column. Mobile phases in each dimension were either methanol or acetonitrile. The sample matrix that was employed in this study contained a mixture of 58 components, comprised of stereoisomers and structural isomers of a systematic variation in molecular weight. Each of the components were oligostyrenes, with between two and five configurational repeating units having either n-butyl, sec-butyl or tert-butyl end groups. In the two-dimensional systems employed in this study, between 46 and 49 of the 58 components could be separated, depending on the mobile phase combinations--with apparent resolution that could not possibly be achieved in a single one-dimensional separation. The results from this study indicate that in order to fully evaluate the resolving power of a 2D system multiple methods of analysis that evaluate the separation potential are most appropriate. This becomes increasingly important when the sample contains components that are very closely related and the retention of solutes displays a high degree of solute crowding.

Molecular Weight↗
PubMed1997

Modification of polystyrenic matrices for the purification of proteins. Effect of the adsorption of poly(vinyl alcohol) on the characteristics of poly(styrene-divinylbenzene) beads for use in affinity chromatography.

A poly(styrene-divinylbenzene) (PSDVB) chromatography matrix, CG1000-sd (TosoHaas), has been modified using poly(vinyl alcohol) (PVA) to create a matrix suitable for the attachment of functional groups for the selective purification of proteins. The characteristics of the modified matrix have been studied using a BET nitrogen adsorption/desorption technique and it has been found that the adsorption of PVA results in the bead micropores being filled whilst the bead macropores are left essentially unaltered. There was no protein adsorption onto the modified matrices. A dye ligand (Procion Blue MX-R) has been covalently attached to PVA-PSDVB matrix and the lysozyme capacities of the PVA-PSDVB matrix have been determined. The matrix compares well with commercial Blue Sepharose Fast Flow, an affinity matrix on cross-linked agarose. The dye-PVA-PSDVB matrix is stable when subjected to sanitisation with sodium hydroxide.

Adsorption↗
PubMed1997

Modification of polystyrenic matrices for the purification of proteins. II. Effect of the degree of glutaraldehyde-poly(vinyl alcohol) crosslinking on various dye ligand chromatography systems.

A poly(styrene-divinylbenzene) chromatography matrix, CG1000sd (TosoHaas) has been modified by the adsorption and crosslinking of poly(vinyl alcohol) (PVA) to create a matrix suitable for the attachment of dye ligands for the adsorption of lysozyme. However, it is shown that there was limited recovery and repeated drops in capacity with adsorption of human serum albumin (HSA). The effect of changing the nature of the PVA crosslinking on the HSA binding characteristics was studied, as well as the effect of using differing dye ligands. The total amount of irreversible HSA binding decreased with greater crosslinking and there were large differences in HSA adsorption characteristics between differing dye types.

Adsorption↗