Aqueous Gel Permeation Research Articles

Molecular weight distribution of chlorolignin in bleached Kraft effluent by GPC and ultrafiltration

The molecular weight distribution (MWD) of chlorinated compounds of bleached Kraft pulp mill effluent was investigated by aqueous gel permeation chromatography (GPC) and ultrafiltration. The effluent was fractionated by ultrafiltration, using different cut-off membranes (MW 50 000, 20 000, 10 000 and 8000). The retentate and permeate of each ultrafiltration was analyzed by GPC. The results showed that the molecular weight distribution for all samples ranged from 200 to 550 Da. Two effects were discussed: associative interactions between the compounds to form high molecular associated complexes (aggregates); and non-size-exclusion effects, like ion exclusion. Aggregates formation was confirmed by ultrafiltration experiments, using a 50 000 cut-off membrane.

Characterization of poly[di(carboxylatophenoxy) phosphazene] by an aqueous gel permeation chromatography

A water-soluble phosphazene polyelectrolyte, poly[di(carboxylatophenoxy)phosphazene] (PCPP), was characterized using aqueous gel-permeation chromatography (GPC) with concentration (UV and RI) and molecular weight sensitive (multiangle laser light-scattering) detectors. Agreement was observed between the weight-average molecular weights determined by GPC with a light-scattering detector, conventional GPC using fractionated narrow PCPP standards, and also by static light-scattering measurements. The effect of chromatography conditions, such as ionic strength of the mobile phase, column resolution, and injection volume was investigated. Mark–Houwink constants of PCPP in aqueous solution (phosphate buffer, pH 7.4, 0.42MNaCl) were determined. The validity of the universal calibration curve and the occurrence of a secondary nonexclusion mechanism of separation in aqueous GPC of phosphazene polyelectrolytes are discussed. © 1996 John Wiley & Sons, Inc.

Characterization of poly[di(carboxylatophenoxy) phosphazene] by an aqueous gel permeation chromatography

A water-soluble phosphazene polyelectrolyte, poly[di(carboxylatophenoxy)phosphazene] (PCPP), was characterized using aqueous gel-permeation chromatography (GPC) with concentration (UV and RI) and molecular weight sensitive (multiangle laser light-scattering) detectors. Agreement was observed between the weight-average molecular weights determined by GPC with a light-scattering detector, conventional GPC using fractionated narrow PCPP standards, and also by static light-scattering measurements. The effect of chromatography conditions, such as ionic strength of the mobile phase, column resolution, and injection volume was investigated. Mark–Houwink constants of PCPP in aqueous solution (phosphate buffer, pH 7.4, 0.42MNaCl) were determined. The validity of the universal calibration curve and the occurrence of a secondary nonexclusion mechanism of separation in aqueous GPC of phosphazene polyelectrolytes are discussed. © 1996 John Wiley & Sons, Inc.

Ion-pair reversed-phase and low-molecular-weight aqueous gel permeation high-performance liquid chromatography methods for the determination of amoxicillin oligomersa

Ion-pair reversed-phase and low-molecular-weight aqueous gel permeation high-performance liquid chromatography methods for the determination of amoxicillin oligomersa

Ion-pair reversed-phase and low-molecular-weight aqueous gel permeation high-performance liquid chromatography methods for the determination of amoxicillin oligomers

Polymeric substances formed from concentrated sodium amoxicillin in an aqueous solution have been separated using two high-performance liquid chromatographic methods. We used a C18 reversed-phase column with tetrabutylammonium chloride as an ion-pairing agent with an acetonitrile gradient and a TSKgel G2500PWxl column with water as the solvent for gel permeation chromatography. The separated materials, ranging in size from the monomer to the tetramer, have been characterized by functional-group chemical analysis, while the identification of the piperazine-2,5-dione was done using a pure standard. A greater number of peaks which were also better defined were obtained using the ion-pair reversed-phase method, and open and closed beta-lactam ring polymer forms could be distinguished. Using the gel permeation method, only a few monomer, piperazine-2,5-dione, dimer, trimer and combined amoxicillin trimer and tetramer peaks were obtained with water, although those obtained were quite well defined. The data on the time-course of formation of the oligomers and the amoxicillin degradation product were virtually identical by both methods.

Degradation of poly( d,l-lactic acid) microspheres: effect of molecular weight

Degradation behaviors of two poly( d,l-lactic acid) (PDLA) microspheres prepared from two different molecular weights (17000 and 41000) were examined. The degree of the degradation was quantitatively estimated by employing an aqueous gel permeation chromatography (GPC) which allowed the determination of the overall amount of water soluble degradation products released out into aqueous medium. At the initial stage of incubation, short chain oligomers, which were produced in an ultrasonication treatment step during the microsphere preparation were immediately released out. Critical weight average molecular weight of the water soluble oligomers ranged from 1050 to 1150. The polymer degradation behaviors of the microspheres during the 53-day incubation were greatly affected depending on the molecular weight of raw polymers as evidenced by significantly different trends of time-dependent molecular weight change. The lower molecular weight polymeric mocrosphere exhibited a significant degradation with reduced glass transition temperature, while the higher molecular weight polymeric microsphere did not show any detectable change in the degradation until 53 days. It was found the water hydration in the low molecular weight PDLA microsphere immediately allowed the polymer morphology to change from a glassy to rubbery state by lowering the glass transition temperature below the incubation temperature. This led to the more susceptible physical state to the degradation. On the other hand, the high molecular weight microsphere was in the glassy state until they started to degrade.

Universal calibration assessment in aqueous gel permeation chromatography

AbstractIt was the objective of this paper to assess the applicability of the universal calibration method to aqueous GPC/SEC with nonionic and anionic polymers using the Viscotek differential viscosity detector. Three water‐soluble polymers—polyacrylic acid, dextran, and polyethylene oxide—were chromatographed using four UltrahydrogelTM columns with 0.3 M NaCl and 0.1M KH2PO4 as the mobile phase adjusted to pH 7. Three distinct calibration curves were obtained. Upon addition of 10% methanol, a reasonably good universal calibration curve was obtained. However, quantitative analysis of the data exhibited about 5% deviation in average Mw and Mn for sodium polyacrylate as calculated from the single curve as opposed to about 40% when calculated from the composite curve. The applicability of three theoretical models for the universal calibration method was assessed, and a recommendation was made for future work. © 1993 John Wiley & Sons, Inc.

Application of Aqueous Gel Permeation Chromatography with In-Line Multi-Angle Laser Light Scattering and Differential Viscometry Detectors for the Characterization of Natural Product Carbohydrate Pharmaceuticals

Abstract Natural product complex carbohydrate polymers are currently being developed as prophylactic and/or therapeutic drugs. These water-soluble carbohydrate pharmaceuticals, which are primarily β-1, 3-D-glucan polymers, belong to the class of drugs known as biological response modifiers (BRMs). A major obstacle to the development, understanding and clinical utilization of carbohydrate BRMs has been the difficulty involved in accurately characterizing high molecular weight (MW) carbohydrate polymers. Recent advances in aqueous gel permeation chromatography (GPC), differential viscometry (DV) and multi- significance of these observations are readily apparent. As stated above, the characterization and quality control of natural product carbohydrate BRMs has, in the past, relied heavily on bioassays and conventional GPC. The data presented demonstrate the applicability of high performance GPC with multiple detectors in the analysis of complex carbohydrate BRMs. This technology when combined with other analytical techniques such as high-field 13C and 1H-NMR, elemental analysis, helix coil transition analysis and exhaustive methylation/gas chromatography/mass spectrometry (12) provide a much more complete description of this novel class of pharmaceuticals.

Development of Macroporous Polymer Packings for Aqueous High Flow Rate Liquid Chromatography. Approach on the Viewpoint of Materials Chemistry

Gel production methods have recently been improved, and a systematic search for packing materials has led to the creation of various hard particles from hydrophilic condensation polymers, making it possible for aqueous gel permeation chromatographic separation to occur at higher flow rates. In the present review, we explain the production methods, efficiency, and applications of hydrophilic packings developed due to this improvement in packing materials, while especially emphasizing their hardness.

Study of the reaction rates and structures of a phenol-formaldehyde resol resin by carbon-13 NMR and gel permeation chromatography

Carbon-13 NMR aqueous gel permeation chromatography studies were carried out on 10 intermediate samples obtained during the synthesis of a typical phenol-formaldehyde resol wood adhesive resin. Apparent kinetic parameters and an improved description of chemical structures in terms of the degree of polymerization and the degree of branching were obtained. The phenol-formaldehyde resol resin of this study was relatively low in molecular weight and high in branches with terminal o,o-bis(hydroxymethyl)phenol groups

Influence of prior exposure to xenobiotics on the metabolism and distribution of polychlorinated biphenyls and phenanthrene in winter flounder, Pseudopleuronectes americanus

The purpose of this study was to determine the effect of prior exposure and multiple-contaminant exposure on the ability of winter flounder to metabolize specific organic compounds. In addition to these pharmacokinetic studies, the intracellular binding patterns of phenanthrene metabolites to liver cytosolic constituents of winter flounder from the different treatments were investigated. In the first experiment, one group of winter flounder was fed by gavage a dose of 14C-labeled polychlorinated biphenyls (PCBs); a second group of fish received a dose of both PCB and cadmium. In a second experiment, winter flounder were exposed for four weeks in a flow-through system to a mixture of copper, cadmium, lead, and PCBs before they received either a dose of 14C-PCB or a dose of 14C-phenanthrene. Liver cytosols from the second experiment were chromatographed by aqueous gel permeation chromatography (AGPC) and the radioactivity associated with 1 ml fractions of the eluate determined by liquid scintillation counting. The results of these experiments suggest that both pre-exposure and/or multiple-contaminant exposures increase the rate at which winter flounder can metabolize both PCBs and phenanthrene. The results also suggest that phenanthrene metabolites were bound to enzymes. There were no apparent differences in the intracellular distribution of radioactivity or the UV and fluorescence chromatograms between liver cytosols from animals from the different treatments.

On-Stream HPLC Determination of Residual Ethylene Oxide and Propylene Oxide in Glycerol Based Polyol Synthesis

Abstract A simple automated HPLC scheme was developed to monitor synthetic polymerization reaction on-stream. Chromatographic conditions that affect the separation of the reaction species from the reaction product and base catalyst using high-performance aqueous gel permeation chromatography were also studied. Reactor samples are automatically drawn from the reaction vessel through a micro-loop sampling valve/actuator assembly and transferred to the chromatographic system via stainless steel tubing without interruption of the mobile phase flow. The resulting chromatograms, monitored by a refractive index detector, of reactants and products are continuously measured at intervals during the reaction automatically and unattended. Application of this system for the reaction of propylene oxide and ethylene oxide to high molecular weight polyols is presented.

Studies on chitosan: 1. Determination of the degree of N-acetylation of chitosan by ultraviolet spectrophotometry and gel permeation chromatography

A new method for determining the degree of N-acetylation of chitosan is proposed, which is based on ultraviolet (u.v.) absorbance at 220 nm of acetamide groups. Using the molar absorption coefficient of the acetamide group of N- acetyl- d-glucosamine and N,N′- diacetyl- d-chitobiose , the degree of N-acetylation of chitosan could be calculated. This principle was applied on high pressure liquid chromatography with an aqueous gel permeation support and a.u.v. detector. It was found that u.v. absorption peak areas in chromatograms were proportional to the concentration of acetamide groups and by this relationship the degree of N-acetylation could be estimated. This method is very useful for the characterization of chitosan because not only the degree of N-acetylation but also molecular weight can be measured at a time. This work indicates that the method is quite useful for tracing the time course of reactions between chitosan and reagents (acetic anhydride and others) because low molecular weight compounds are swept out from the main peak of chitosan in the chromatogram and the separation and purification of a reaction product can be omitted.

Small-angle x-ray scattering of high- and low-affinity heparin

Solution characterization of heparin with high affinity (HA) and low affinity (LA) for antithrombin III was performed using the methods of small-angle x-ray scattering (SAXS), viscometry, and aqueous gel permeation chromatography (GPC). SAXS provided various topological parameters including the radius of gyration ( 〈S 2〉 1 2 ), radius of gyration of cross-section ( 〈S 2〉 q 1 2 ), persistence length ( a ∗ ), contour length ( L), and mass parameters e.g., overall molecular mass ( M r), and mass per unit length ( M q). The molecular weights of HA and LA pig mucosal heparins were found to be 14,900 and 11,500 and the respective radii of gyration were 40.1 and 33.6 Å. The persistence lengths of HA and LA were 21.3 and 20.3 Å, respectively. These parameters were compared to SAXS data of heparin [S. S. Stivala, M. Herbst, O. Kratky, and I. Pilz (1968) Arch. Biochem. Biophys. 127, 795–802] fractionated according to molecular weight only. It was found that the various experimental values of this heparin lie somewhere in between those of HA and LA heparins. It appears that there are no appreciable differences in the physico-chemical properties, including conformation, among the heparins in H 2O at 25 °C.

High-speed aqueous gel permeation chromatography using a poly(vinyl alcohol) hollow fibre

High-speed aqueous gel permeation chromatography using a poly(vinyl alcohol) hollow fibre

System development for aqueous gel permeation chromatography

AbstractTo devise an improved gel permeation chromatography system for the analysis of macromolecules, with a strongly alkaline solution as eluent, three Fractogel TSK gels were investigated. Because primary standards are usually unavailable, calibration was achieved by using Benoit's universal procedure and polystyrene sulfonates, polyethylene glycols, and dextrans as secondary calibrants. Continuous monitoring of intrinsic viscosity resulted from coupling a viscodetector with a classical differential refractometer. It was found that column efficiency and fractionation range could be extended well beyond those of previously studied systems on a reasonably short period of analysis.

Determination of zinc and zinc-containing compound in rabbit aqueous humour by high performance aqueous gel permeation chromatography-atomic absorption spectrophotometry.

Zinc content in rabbit aqueous humour was first determined by atomic absorption spectrophotometry. In the normal rabbit, the content ranged from 0.04 to 0.06 mg/l, but in the rabbit with steroid-induced glaucoma ( SIG ), it increased and ranged from 0.1 to 0.50 mg/l. Further, in the steroid ( cortisone, 0.3 mg ) injected rabbit ( SIR ), the zinc content ranged from 0.06 to 0.09 mg/l. The precision of the measurement was within 0.01 mg/l. The minimal detection limit was 0.03 mg/l. A zinc-containing compound ( Mw, about 6000 ) was detected in the aqueous humour with the combined system of a high performance aqueous gel permeation chromatograph and an atomic absorption spectrophotometer. This zinc compound was always induced in the aqueous humour when the total zinc content exceeded 0.1 mg/l, i.e., in the case of the rabbit with SIG. On the other hand, the zinc compound could be occasionally induced in the fluid of SIR but not in the normal one. The analytical time required for the detection of the zinc compound was within 40 min. The usefulness of the analysis of zinc and the zinc-containing compound in the aqueous humour is suggested for the clinical diagnosis of SIG, one of the ophthalmic diseases.

Light scattering detection in aqueous high-performance gel permeation chromatography (HPGPC): Relative molecular mass distribution of eye lens proteins

Light scattering techniques have been greatly improved since laser light sources, enabling dynamic light scattering and measurements at low angles have been available. Relative molecular mass determinations can be performed at low solute concentration, extrapolations to zero-angle are no longer required and, because of the low scattering volume, a flow-through cell construction has become possible where dust particles only interfere as spikes [1]. For HPLC detection the low-angle laser light scattering (LALLS) signal corresponds to the product of relative molecular mass and concentration; combination with a concentration detector (for example, differential refractometer, UV absorbance) enables calculation of relative molecular mass [2]. The sensitivities of LALLS and the usual concentration detectors are comparable for (bio)polymers with relative molecular masses near 100,000. The minimum relative molecular mass to be determined in this way is near 10,000 and, therefore, the only mode of HPLC using the LALLS detector to date is gel permeation (size exclusion) chromatography.

Urinary protein profiling by high-performance gel permeation chromatography

We describe a new application of high-performance aqueous gel permeation chromatography for the analysis of human proteinuria. Separations of urinary proteins from normal subjects and patients with renal impairment were performed with TSK G 3000 SW columns. The effects of pH and icnic strength of the eluent on the separation of urinary proteins were investigated. Albumins were selectively separated from urine by affinity chromatography on Blue Sepharose CL-6B. According to the results of clinical investigations, urinary protein pattern derived from gel permeation chromatography revealed a good prediction of the site of renal involvement. Predominant excretion of proteins with lower molecular weight than albumin correlated with tubular damage. Albumin and higher molecular weight protein patterns were associated with glomerular disease. Absorbance measurements of the eluent at 280 nm were used for quantitative determination of total urinary protein. Gel permeation chromatography was compared to sodium dodecyl sulfate—polyacrylamide gel electrophoresis and the resulting protein patterns are in good agreement.

Molecular weight distributions of cationic polymers by aqueous gel permeation chromatography

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