| 21. |
- Gustavsson, Per-Erik, et al.
(author)
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Superporous agarose, a new material for chromatography
- 1996
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In: Journal of chromatography. A. - : Elsevier BV. - 0021-9673. ; 734:2, s. 231-240
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Journal article (peer-reviewed)abstract
- This paper reports on a new type of spherical agarose chromatography particles characterized by two sets of pores, normal diffusion pores, characteristic of all agarose materials and very wide pores, so-called superpores or flow pores. These superpores allow part of the chromatographic flow to pass through each individual particle, which gives improved mass transfer, especially in situations where diffusion is the limiting factor for the overall performance of a chromatographic separation. The particles were prepared by a double emulsification procedure. Observations under a microscope and size-exclusion chromatography were used in order to demonstrate pore flow. The chromatographic behaviour of the new particles was as efficient as that of homogeneous particles which were several times smaller. The agarose particles were derivatized with polyethyleneimine and used for an ion-exchange chromatographic separation of three model proteins. As expected from a perfusion material, the superporous beads resolved the protein mixture more efficiently than homogeneous beads of the same size.
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| 22. |
- Gustavsson, Per-Erik, et al.
(author)
-
Superporous agarose as an affinity chromatography support
- 1997
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In: Journal of chromatography. A. - 0021-9673. ; 776:2, s. 197-203
-
Journal article (peer-reviewed)abstract
- Superporous agarose beads were used as an affinity support in column chromatography. These beads characteristically possess two sets of pores, normal diffusion pores and flow pores, so-called superpores. The superpores, whose diameter is a substantial fraction of the particle diameter (i.e. 1/3 to 1/10 of the particle diameter), allow part of the chromatographic flow to pass through each individual bead. Consequently, significant improvement in mass transfer is observed in superporous beads as compared with homogeneous beads, especially at high flow-rates [Gustavsson and Larsson, J. Chromatogr. A, 734 (1996) 231–240.]Superporous agarose beads and homogeneous agarose beads were each derivatized with two types of affinity ligands. A NAD+ analogue was used for the purification of bovine lactate dehydrogenase and protein A was used for the adsorption of rabbit IgG. The performances of superporous beads and homogeneous beads were compared. Superporous bead columns derivatized with protein A and NAD+ analogue could be operated 5 times and 3 times, respectively, as fast as corresponding homogeneous bead columns.
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| 23. |
- Gustavsson, Per-Erik, et al.
(author)
-
Superporous agarose beads as a hydrophobic interaction chromatography support
- 1999
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In: Journal of Chromatography A. - 0021-9673. ; 830:2, s. 275-284
-
Journal article (peer-reviewed)abstract
- Superporous agarose beads were used as a support for hydrophobic interaction chromatography. These beads have large connecting flow pores in addition to their normal diffusion pores. The flow pores, which are approximately one fifth of the overall diameter of the superporous agarose beads, were earlier shown to give the beads improved mass transfer properties relative to homogeneous agarose beads (Gustavsson and Larsson, J. Chromatogr. A, 734 (1996) 231-240). Superporous agarose beads and homogeneous agarose beads of the same particle size range (106-180 mu m) were derivatized with phenyl groups. The properties of the superporous beads were then compared with the homogeneous beads in the separation of a mixture of three model proteins (ribonuclease A, lysozyme and bovine serum albumin) at various superficial flow velocities from 30 to 600 cm/h. The superporous beads gave satisfactory separation at flow velocities five times higher than was possible for homogeneous beads. The performance of the two types of beads was also compared in the purification of lactate dehydrogenase from a beef heart extract at a superficial flow velocity of 150 cm/h. The superporous beads performed considerably better, leading to twice the purification factor and twice the concentration of the desired product. The results were interpreted using the theoretical treatment given by Carta and Rodrigues (Carta and Rodrigues, Chem. Eng. Sci., 48 (1993) 3927). (C) 1999 Elsevier Science B.V. All rights reserved.
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| 24. |
- Jeppsson, Jan olof, et al.
(author)
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Isolation and characterization of two minor fractions of α1,-antitrypsin by high-performance liquid chromatographic chromatofocusing
- 1985
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In: Journal of Chromatography A. - 0021-9673. ; 327, s. 173-177
-
Journal article (peer-reviewed)abstract
- α1-Antitrypsin is a glycoprotein that separates into five electrophoretic fractions, viz. M2, M4, M6, M7 and M8. Con A-Sepharose separates the protein into three fractions according to the branching degree of the three oligosaccharide chains. The Con A affinity is identical for M4 and M7 and for M6 and M8. Within each pair the proteins were isolated by rapid chromatofocusing. The M7 and M8 have the same carbohydrate structure as the major M4 and M6 respectively, but have lost the first five N-terminal amino acids (Glu-Asp-Pro-Glu-Gly) as compared to the majority of the protein.
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| 25. |
- Jönsson, Malin, et al.
(author)
-
Protein partitioning in thermoseparating systems of a charged hydrophobically modified ethylene oxide polymer
- 2003
-
In: Journal of Chromatography A. - 0021-9673. ; 983:1-2, s. 133-144
-
Journal article (peer-reviewed)abstract
- The phase behavior of a thermoseparating cationic hydrophobically modified ethylene oxide polymer (HM-EO) containing tertiary amines has been investigated at different pH, salt and sodium dodecyl sulfate (SDS) concentrations, in order to find a water/HM-EO two-phase, system suitable for protein partitioning. The used polymer forms micellar aggregates that can be charged. By changing pH and SDS concentrations the netcharge of the SDS/HM-EO aggregate can be shifted from positive to negative. Bovine serum albumin (BSA) and lysozyme were partitioned in the thermoseparated two-phase systems of the cationic polymer at different pH, salt and SDS concentrations. The dominant attractive interactions between the polymer aggregates and the studied proteins were shown to be of electrostatic (Coulomb) nature rather than hydrophobic interaction. At low ionic strength the positively charged polymeric aggregates attracted negatively charged BSA and repelled positively charged lysozyme. Upon addition of SDS the negatively charged aggregates attracted lysozyme and repelled BSA. Thus, it was possible to direct proteins with different charges to the polymeric phase and redirect them to a polymer-depleted phase by changing the netcharge of the polymeric aggregates. The effect of different salts on the partitioning of BSA in a system of slightly positively charged HM-EO was studied. NaCl and KBr have a significant effect on driving the BSA to the polymer-depleted phase, whereas KF and K2SO4 have a smaller effect on the partitioning. The cloud point temperature of the charged polymer decreased upon addition of SDS near the isoelectric molar ratio of SDS to polymer and also upon salt addition. In the latter case the decrease was smaller than expected from model calculations based on Flory-Huggins theory, which were performed for a charged thermoseparating polymer at different charges and salt concentrations. (C) 2002 Elsevier Science B.V. All rights reserved.
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| 26. |
- Kagedal, Bertil, et al.
(author)
-
Automated high-performance liquid chromatographic determination of 5-S-cysteinyl-3,4-dihydroxyphenylalanine in urine
- 1989
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In: Journal of Chromatography A. - 0021-9673. ; 473, s. 359-370
-
Journal article (peer-reviewed)abstract
- An automated high-performance liquid chromatographic (HPLC) method has been developed for measurement of 5-S-cysteinyl-DOPA in urine (DOPA = 3,4-dihydroxyphenylalanine). The urinary sample was injected into an HPLC boronate column. With a mobile phase of 0.1 M phosphate buffer containing 0.2 mM disodium ethylenediaminetetraacetate (Na2EDTA) (pH 6.0) mixed with methanol (9:1), 5-S-cysteinyl-DOPA was adsorbed while most other compounds were washed away. By column switching, the column flow was reversed and 5-S-cysteinyl-DOPA was desorbed by a mobile phase of 0.1 M formic acid and 0.2 mM Na2EDTA at pH 3.0 and chromatographed on a reversed-phase column. The precision, as estimated from repeated analysis of an urinary sample and from duplicate analysis of a number of samples, ranged from 1.4 to 5.2% (coefficient of variation), and the analytical recovery was 93 +/- 4.1%. The method is suitable for use in the clinical laboratory.
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| 27. |
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| 28. |
- Kempe, Henrik, et al.
(author)
-
Simulation of chromatographic processes applied to separation of proteins
- 1999
-
In: Journal of Chromatography A. - 0021-9673. ; 846:1-2, s. 1-12
-
Journal article (peer-reviewed)abstract
- The advantages of using a detailed mathematical model for fixed bed chromatography is demonstrated by the personal computer program SIMCHROM. The chromatography model includes axial dispersion in the bulk liquid, external and internal mass transfer resistances and an instationary non-linear adsorption model. Frontal and pulse chromatography can be studied for single and multicomponent systems. The simulation program can easily be used to make parametric evaluations to study the influence of variations in physical, kinetical and operating parameters. The special features of the present intrinsic model is demonstrated by comparing the SIMCHROM results With simulations using simplified lumped models. Experimental data describing affinity chromatography of lysozyme on Cibacron Blue Sepharose CL-6B is used as a model system. The intrinsic model is able to describe variations in the physical, kinetic and operating parameters better than the simplified models. This results in a more reliable prediction of the performance of the chromatography process as well as a better understanding of che underlying mechanisms responsible for the separation. (C) 1999 Elsevier Science B.V. All rights reserved.
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| 29. |
- Kempe, Maria, et al.
(author)
-
Direct resolution of naproxen on a non-covalently molecularly imprinted chiral stationary phase
- 1994
-
In: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 664:2, s. 276-279
-
Journal article (peer-reviewed)abstract
- A synthetic polymer selective for (S)-naproxen was prepared by molecular imprinting. 4-Vinylpyridine and ethylene glycol dimethacrylate were copolymerised in the presence of the template, (S)-naproxen. The template was extracted from the polymer, leaving specific recognition sites, complementary to the template. The polymer was utilized as a stationary phase in HPLC. Racemic naproxen was efficiently resolved on the polymer. Furthermore, the polymer was able to separate naproxen from the structurally related ibuprofen and ketoprofen.
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| 30. |
- Kempe, Maria, et al.
(author)
-
Molecular imprinting used for chiral separations
- 1995
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In: Journal of Chromatography A. - 0021-9673. ; 694:1, s. 3-13
-
Research review (peer-reviewed)abstract
- Molecular imprinting is a promising technique for the preparation of synthetic polymers of predetermined specificity. Functional monomers are copolymerized with crosslinkers in the presence of the desired molecule, the imprint molecule. The use of these polymers as chiral stationary phases is discussed. Other applications, such as antibody-mimics, enzyme-like catalysts and sensors, are also focused upon.
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