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  • 1.
    Amini, Ahmad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Separation of somatropin charge variants by multiple-injection CZE with Polybrene/chondroitin sulfate A double-coated capillaries2013In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 36, no 16, p. 2686-2690Article in journal (Refereed)
    Abstract [en]

    The performance of dynamic double-coated fused-silica capillaries with Polybrene and chondroitin sulfate A has been compared with uncoated fused-silica capillaries for the determination of recombinant human growth factor (somatropin) charge variants. The separations were carried out under the same electrophoretic conditions as described in the European Pharmacopoeia, i.e. at pH 6.0 and 30 degrees C. The coating significantly reduced the interactions between the proteins and the surface of the fused-silica capillary. The first five separations performed in a new bare fused-silica capillary were discarded because of very poor separation performance as a result of protein-surface interactions. There was an approximate twofold increase in the interday migration time precision (%RSD 6.5%) in the double-coated capillaries. The method was successfully transferred to a multiple CZE mode where two samples were analyzed in a single electrophoretic run. The average purity of somatropin certified reference standard was 98.0% (%RSD 0.3%) determined by using uncoated and coated capillaries.

  • 2. Bertucci, Carlo
    et al.
    Pistolozzi, Marco
    Felix, Guy
    Danielson, Helena
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    HSA binding of HIV protease inhibitors: a high-performance affinity chromatography study2009In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 32, no 10, p. 1625-1631Article in journal (Refereed)
    Abstract [en]

    The binding of HIV protease inhibitors, drugs important for anti-HIV chemotherapy, to HSA was examined by high-performance affinity chromatography. Frontal analysis was first used to determine the amount of anchored protein and the binding capacity for selected markers on this column. Zonal elution experiments then ranked the HSA bound fraction of the examined compounds. Information on the g region was obtained by competitive zonal elution experiments using probe binding compounds with known sites on HSA. An allosteric competition between HIV protease inhibitors (PIs) and valproate (a probe for the bilirubin site) was detected, consistent with a noncooperative binding mechanism. No significant competition was observed between the examined compounds and salicylate or ibuprofen, probes for sites I and II, respectively. The observations were confirmed by circular dichroism spectroscopy, based on the change in the induced circular dichroism signals of selected markers for the main binding sites of HSA when ritonavir was added as the competitor. These results were in good agreement with previous literature reports and provide more details on how PIs are transported in plasma and how they may compete with other drugs in the body.

  • 3. Gao, Min
    et al.
    Wang, Xiaolei
    Gu, Ming
    Su, Zhiguo
    Wang, Ye
    Janson, Jan-Christer
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Separation of polyphenols using porous polyamide resin and assessment of mechanism of retention2011In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 34, no 15, p. 1853-1858Article in journal (Refereed)
    Abstract [en]

    A porous polyamide resin is shown to possess hydrogen bond acceptor properties suitable for the separation of polyphenolic solutes such as phenolic acids, flavonols and flavonoids. The separation is achieved in the presence of solvent mixtures of acetic acid and ethanol. The extent of hydrogen bond adsorption is reviewed based on data obtained from the elution behaviour of a variety of simple polyphenolic solutes. Polyamide adsorption chromatography was applied for the purification of resveratrol and polydatin from Polygonum cuspidatum Sieb. & Zucc.

  • 4.
    Ghasemzadeh, Nasim
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Nyberg, Fred
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Hjertén, Stellan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Highly selective artificial gel antibodies for detection and quantification of biomarkers in clinical samples: I. Spectrophotometric approach to design the calibration curve for the quantification2008In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 31, no 22, p. 3945-3953Article in journal (Refereed)
    Abstract [en]

    High selectivity of a biomarker is a basic requirement when it is used for diagnosis, prognosis and treatment of a disease. The artificial gel antibodies, which we synthesise by a molecular imprinting method, have this property not only for proteins, but also for bioparticles, such as viruses and bacteria. However, diagnosis of a disease requires not only that the biomarker can be "fished out" from a body fluid with high selectivity, but also that its concentration in the sample can rapidly be determined and preferably by a simple technique. This paper deals primarily with the development of a spectrophotometric method, which is so simple and fast that it can be used with advantage in a Doctor's Office. The development of this method was not straight-forward. However, by modifications of the performance of these measurements we can now design standard curves in the form of a straight line, when we plot the true (not the recorded "apparent" absorption) against known protein concentrations. In an additional publication (see the following paper in this issue of JSS) we show an application of such a plot: determination of the concentration of albumin in serum and cerebrospinal fluid from patients with neurological disorders to investigate whether albumin is a biomarker for these diseases.

  • 5.
    Ghasemzadeh, Nasim
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Nyberg, Fred
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Hjertén, Stellan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Highly selective artificial gel antibodies for detection and quantification of biomarkers in clinical samples: II. Albumin in body fluids of patients with neurological disorders2008In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 31, no 22, p. 3954-3958Article in journal (Refereed)
    Abstract [en]

    We have previously used the molecular-imprinting method for the synthesis of artificial gel antibodies, highly selective for various proteins. In the present work, we have synthesized artificial gel antibodies against human albumin with the aim to develop a simple and rapid procedure to measure the concentration of this protein in samples of clinical interest. The procedure, based on the design of a standard curve (see the preceding paper), was applied on a quantitative analysis of albumin in human plasma and cerebrospinal fluid (CSF). We found that our technique permitted detection of albumin in these body fluids with high precision and that the concentration of this protein was significantly enhanced in CSF from patients with amyotrophic lateral sclerosis (ALS), compared to control samples. This finding is in agreement with results from earlier studies, which confirms the validity of our analysis technique and suggests that the barrier permeability may be affected in ALS, perhaps also for other proteins. No enhancement in plasma levels of albumin was seen in patients with ALS, but rather a decrease. The results further indicate that our approach might also apply well to other biomarkers for the actual neurological disease and other disorders.

  • 6.
    Jansson, Erik T.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Strategies for analysis of isomeric peptides2018In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 41, no 1, p. 385-397Article, review/survey (Refereed)
    Abstract [en]

    This review presents an overview and recent progress of strategies for detecting isomerism in peptides, with focus on D/L epimerization and the various isomers that the presence of an aspartic acid residue may yield in a protein or peptide. While mass spectrometry has become a majorly used method of choice within proteomics, isomerism is inherently difficult to analyze because it is a modification that does not yield any change in mass of the analyte. Here, several techniques used for analysis of peptide isomerism are discussed, including enzymatic assays, liquid chromatography, and capillary electrophoresis. Recent progress in method development using mass spectrometry is also discussed, including labeling strategies, fragmentation techniques, and ion-mobility spectrometry.

  • 7. Liu, Dan
    et al.
    Ma, Yan
    Gu, Ming
    Janson, Jan-Christer
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Wang, Changhai
    Xiao, Hongbin
    Liquid-liquid/solid three-phase high-speed counter-current chromatography, a new technique for separation of polyphenols from Geranium wilfordii Maxim2012In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 35, no 16, p. 2146-2151Article in journal (Refereed)
    Abstract [en]

    High-speed counter-current chromatography using a new liquidliquid/solid three-phase system was used for the separation of the polyphenols corilagin and geraniin from a crude extract of Geranium wilfordii Maxim in one step. The optimized three-phase system was composed of n-hexane/ethyl acetate/methanol/acetic acid/water and to which was added 10-mu m average diameter microspheres of cross-linked 12% agarose at the ratio of 0.2:10:2:1:5 and 0.1 g/mL, respectively. The purities of geraniin and corilagin were 82 and 90%, which were determined by HPLC at 280 nm. A 14.5 and 7 mg of geraniin and corilagin were purified from 160 mg crude extract with the yields of 70 and 78%, respectively.

  • 8. Liu, Dan
    et al.
    Ma, Yan
    Wang, Ye
    Su, Zhiguo
    Gu, Ming
    Janson, Jan-Christer
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    One-step separation and purification of hydrolysable tannins from Geranium wilfordii Maxim by adsorption chromatography on cross-linked 12% agarose gel2011In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 34, no 9, p. 995-998Article in journal (Refereed)
    Abstract [en]

    The hydrolysable tannins corilagin and geraniin, the major active components of the traditional Chinese medicine Geranium wilfordii Maxim, have been separated and purified from crude extracts in one step by adsorption chromatography on cross-linked 12% agarose gel (Superose 12 10/300 GL). The separation was achieved by gradient elution using mobile phase A composed of 5% ethanol and 5% acetic acid and mobile phase B composed of 30% ethanol and 30% acetic acid. The gradients were composed as follows: 0-240 mL, 0-25% B; 240-480 mL, 25-40% B; after 480 mL, 100% B. The purities of the collected corilagin and geraniin were 92.4 and 87.2%, and the corresponding yields were 88.0 and 76.8%, respectively.

  • 9.
    Paul, Prasanta
    et al.
    Univ Leuven, KU Leuven, Dept Pharmaceut & Pharmacol Sci, Leuven, Belgium..
    Duchateau, Tom
    Univ Leuven, KU Leuven, Dept Pharmaceut & Pharmacol Sci, Leuven, Belgium..
    Sänger - van de Griend, Cari
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry. Kantisto BV, Baarn, Netherlands..
    Adams, Erwin
    Univ Leuven, KU Leuven, Dept Pharmaceut & Pharmacol Sci, Leuven, Belgium..
    Van Schepdael, Ann
    Univ Leuven, KU Leuven, Dept Pharmaceut & Pharmacol Sci, Leuven, Belgium..
    Capillary electrophoresis with capacitively coupled contactless conductivity detection method development and validation for the determination of azithromycin, clarithromycin, and clindamycin2017In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 40, no 17, p. 3535-3544Article in journal (Refereed)
    Abstract [en]

    A capillary electrophoresis with capacitively coupled contactless conductivity detection based method for the assay of azithromycin, clarithromycin and clindamycin was optimized and validated in this study. A buffer solution of 20 mM 2-(N-morpholino) ethane sulfonic acid, 40 mM L-histidine and 0.6 mM cetyltrimethylammonium bromide (pH 6.39) was used for the electrophoresis. An uncoated, bare-fused silica capillary (total length 60 cm, effective length 32 cm, 75 mu m id) was used at 25 degrees C. The sample was injected hydrodynamically at 0.5 psi for 5 s. The electrophoresis was conducted at 30 kV in reverse polarity for 6 min with 3 and 2 min of in-between sodium hydroxide (0.1 M) and background electrolyte rinsing, respectively. Ammonium acetate was used as internal standard. This simple and robust method showed reasonable limit of detection and limit of quantitation for azithromycin (0.0125/0.03 mg/mL), clarithromycin (0.017/0.03 mg/mL), and clindamycin (0.038/0.06 mg/mL), with good selectivity, precision both intraday (relative standard deviation <= 1.0%) and interday (relative standard deviation < 3.7%), linearity (R-2 > 0.999) and recovery (99 - 101.7%). The method was successfully applied for the determination of azithromycin, clarithromycin and clindamycin in formulations.

  • 10. Samuelsson, Jorgen
    et al.
    Cavazzini, Alberto
    Shalliker, Ross Andrew
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC.
    Regeneration of a silica monolithic rod column using harsh methods followed by firm thermodynamic and kinetic validation2014In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 37, no 8, p. 906-911Article in journal (Refereed)
    Abstract [en]

    In this study, a numerical tool is introducedbased on thermodynamic and kinetic separation theoryfor validating the regeneration of monolithic rod columns after cutting their inlet sections. A long-used RP-18e monolithic column was deemed to be unfit for further coffee analysis because of poor separation performance. The columns brownish inlet section was physically removed with a lathe, leaving a clean white inlet section. The original and regenerated columns were extensively analyzed and compared using numerical tools for processing adsorption data. The perturbation peak method was used to measure the adsorption isotherm of phenol on the original and regenerated monolith and the adsorption energy distributions were calculated for identifying any change in the degree of heterogeneity. Although peak shapes improved considerably after regeneration, no significant differences were found in the detailed characterization of the processed adsorption data between the original column and the regenerated one. This indicates that the removal of a section of the monolithic bed can be undertaken without damaging the column and columns in which their inlet head sections are contaminated may still function with normal adsorption behavior. In addition, the combined thermodynamic and kinetic methodology could accurately be used to evaluate any regeneration method of columns.

  • 11.
    Samuelsson, Jörgen
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Arnell, Robert
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Potential of adsorption isotherm measurements for closer elucidating of binding in chiral liquid chromatographic phase systems2009In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 32, no 10, p. 1491-1506Article, review/survey (Refereed)
    Abstract [en]

    The human body is a chiral environment and many drugs are chiral and interact differently depending on the type of enantiomer. Therefore, the interest in analytical and preparative separations of enantiomers has steadily increased over the years. LC is today the most important technique in analytical laboratories worldwide. The key to understand the separation system lies in the adsorption isotherm, which describes the equilibrium distribution of solutes between the mobile and stationary phases. By measuring adsorption isotherms in chiral phase systems, a deeper interpenetration concerning enantioselective and non-selective binding energies and adsorption processes is possible. Furthermore, this data provides the core information needed to optimize preparative chromatographic processes for purification of single enantiomers. However, the measurement of adsorption isotherms is a delicate matter and there are many dangerous pitfalls that may produce erroneous results and even wrong mechanistic conclusions. This review summarizes the most relevant methods and a workflow will be given for avoiding the common pitfalls and obtaining reliable data. Several applications from the literature are also treated to give insight in what information can potentially be obtained from using this methodology.

  • 12. Takatsy, Aniko
    et al.
    Kilár, Aniko
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Biochemistry.
    Kilár, Ferenc
    Hjertén, Stellan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Biochemistry.
    Universal method for synthesis of artificial gel antibodies by the imprinting approach combined with a unique electrophoresis technique for detection of minute structural differences of proteins, viruses and cells (bacteria): Ia. Gel antibodies against proteins (transferrins)2006In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 29, no 18, p. 2802-2809Article in journal (Refereed)
    Abstract [en]

    Artificial antibodies in the form of gel granules were prepared by the molecular imprinting technique from the monomers acrylamide and N,N-methylene-bis-acrylamide. Gel granules, freed from the selectively adsorbed protein (the antigen), are neutral and, accordingly, do not migrate in an electrical field. However, upon selective interaction with the antigen at a pH different from its pI, the granules become charged. The selectivity of the gel antibodies was studied by free zone electrophoresis in a tube with inside diameter larger than the size of the granules. Such electrophoretic analyses showed that gel antibodies against iron-free transferrin had a high selectivity for this protein, although some crossreaction took place with iron-saturated transferrin, indicating that these artificial antibodies can easily distinguish the minute differences in the 3-D structure of the transferrins. Analogously, gel antibodies against iron-saturated transferrin were highly selective for this protein with some crossreaction with iron-free transferrin. The mobilities of iron-free and iron-saturated transferrin are very similar, and, therefore, capillary free zone electrophoresis cannot distinguish between these structurally related proteins. However, significant differences in the mobilities of the selective gel granules can be observed depending on their interaction with iron-free or iron-saturated transferrin, i.e., the artificial gel antibodies may become powerful analytical tools.

  • 13. Takatsy, Aniko
    et al.
    Sedzik, Jan
    Kilar, Ferenc
    Hjertén, Stellan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Biochemistry.
    Universal method for synthesis of artificial gel antibodies by the imprinting approach combined with a unique electrophoresis technique for detection of minute structural differences of proteins, viruses, and cells (bacteria): II. Gel antibodies against virus (Semliki Forest Virus)2006In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 29, no 18, p. 2810-2815Article in journal (Refereed)
    Abstract [en]

    Artificial and highly selective antibodies (in the form of gel granules) against proteins can easily be synthesized by a simple, cost-effective imprinting technique [Liao, J.-L. et al., Chromatographia 1996, 42, 259-262]. Using the same method for synthesis of gel antibodies against viruses in combination with analysis by free zone electrophoresis in a rotating narrow bore tube we have shown that artificial gel antibodies against Semliki Forest Virus (wild type) can sense the difference between this virus and a mutant, although they differ in their chemical composition only by three amino acids in one of the three proteins on the surface of the virus particle. The reason for this extremely high resolution is explained by the fact that we use three types of selectivity: (i) shape selectivity (created by the close fit between the antigen and its imprint in the gel), (ii) bond selectivity in the contact area between the antigen and its imprint in the gel antibody, and (iii) charge selectivity, originating from slightly different structures or/and conformations of the antigens.

  • 14.
    Undin, Torgny
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Samuelsson, Jörgen
    Department of Engineering and Chemical Sciences, Karlstad University, Karlstad, Sweden.
    Törncrona, Anders
    AkzoNobel Pulp & Performance Chemicals AB, Bohus, Sweden.
    Fornstedt, Torgny
    Department of Engineering and Chemical Sciences, Karlstad University, Karlstad, Sweden.
    Evaluation of a combined linear–nonlinear approach for column characterization using modern alkaline-stable columns as model2013In: Journal of Separation Science, ISSN 1615-9306, E-ISSN 1615-9314, Vol. 36, no 11, p. 1753-1761Article in journal (Refereed)
    Abstract [en]

    This study investigates if deeper understanding is achieved when combining nonlinear and linear chromatographic column characterization methods. As test systems, two hybrid columns (Phenomenex Gemini-NX C18 and Kromasil Eternity C18) and one classic one (Kromasil-C18) were selected. The nonlinear methods were based on firm adsorption theory and involved determination of adsorption isotherms followed by calculations with a new numerical tool, adsorption energy distribution, on probe components at different pH values. The linear methods involved the hydrophobic subtraction model and selected probe components retention factors as a function of pH. The combined analysis indicated that both complementary and confirmative information can be achieved regarding the actual model systems.

1 - 14 of 14
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