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  • 1.
    Agmo Hernández, Víctor
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Samuelsson, Jörgen
    Department of Engineering and Chemical Sciences, Karlstad University, SE-651 88 Karlstad, Sweden.
    Forssén, Patrik
    Department of Engineering and Chemical Sciences, Karlstad University, SE-651 88 Karlstad, Sweden.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry. Department of Engineering and Chemical Sciences, Karlstad University, SE-651 88 Karlstad, Sweden.
    Enhanced interpretation of adsorption data generated by liquid chromatography and by modern biosensors2013In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1317, no SI, p. 22-31Article in journal (Refereed)
    Abstract [en]

    In this study we demonstrate the importance of proper data processing in adsorption isotherm estimations. This was done by investigating and reprocessing data from five cases on two closely related platforms: liquid chromatography (LC) and biosensors. The previously acquired adsorption data were reevaluated and reprocessed using a three-step numerical procedure: (i) preprocessing of adsorption data, (ii) adsorption data analysis and (iii) final rival model fit. For each case, we will discuss what we really measure and what additional information can be obtained by numerical processing of the data. These cases clearly demonstrate that numerical processing of LC and biosensor data can be used to gain deeper understanding of molecular interactions with adsorption media. This is important because adsorption data, especially from biosensors, is often processed using old and simplified methods.

  • 2.
    Amirkhani, Ardeshir
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Analytical Chemistry.
    Wetterhall, Magnus
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Analytical Chemistry.
    Nilsson, Stefan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Analytical Chemistry.
    Danielsson, Rolf
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Analytical Chemistry.
    Comparison between different sheathless electrospray emitter configurations regarding the performance of nanoscale liquid chromatography time-of-flight mass spectrometry analysis2004In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1033, no 2, p. 257-266Article in journal (Refereed)
    Abstract [en]

    Four different sheathless electrospray ionization (ESI) configurations were investigated for a nano liquid chromatography (LC) system. The studied configurations were: a column with an integrated emitter, with the ESI potential applied before or after the column, and a column with separate emitter, with the ESI voltage applied at a union before the emitter or at the emitter tip. The results indicates that the efficiency of the LC system is rather independent of the configuration when using 95 μm i.d. columns, acetic mobile phase and standard peptides as a sample. Introduction of post column dead volume seems not to be a critical issue at least with flow rates down to 600 nl/min.

  • 3.
    Arnell, Robert
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Forssén, Patrik
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Accurate and rapid estimation of adsorption isotherms in liquid chromatography using the inverse method on plateaus2005In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1099, p. 167-174Article in journal (Refereed)
  • 4. Arnell, Robert
    et al.
    Forssén, Patrik
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Sardella, Roccaldo
    Laemmerhofer, Michael
    Lindner, Wolfgang
    Adsorption behaviour of a quinidine carbamate-based chiral stationary phase: Role of the additive2009In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1216, no 16, p. 3480-3487Article in journal (Refereed)
    Abstract [en]

    In this study, we incorporate the additive properties into the theoretical model of a general preparative chromatographic system; this is normally not done and this limits a proper process optimization. As a model phase system, we used the adsorption of 9H-fluoren-9-ylmethoxycarbonyl-allylglycine (Fmoc-allylglycine) enantiomers on a quinidine carbamate-based chiral stationary phase (anion exchanger) together with a methanol-glacial acetic acid-ammonium acetate eluent. The inverse method was used to measure the competitive adsorption isotherms of both the Fmoc-allylglycine enantiomers as well as the non-detectable additive acetic acid. It was concluded that this enantioselective preparative system is well described by a non-heterogeneous adsorption model and that the loading capacity is very high. The proposed model is valid over a wide range of additive concentrations, which is important for process optimization.

  • 5.
    Arvidsson, Björn
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Johannesson, Nina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Citterio, Attilio
    Righetti, Pier Giorgio
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    High throughput analysis of tryptophan metabolites in a complex matrix using capillary electrophoresis coupled to time-of-flight mass spectrometry2007In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1159, no 1-2, p. 154-158Article in journal (Refereed)
    Abstract [en]

    A capillary electrophoresis method for separation and detection with time-of-flight mass spectrometry is described for tryptophan metabolites in the kynurenic pathway. Tryptophan metabolites are usually difficult to detect with electrospray mass spectrometry since they have low surface activity and occur in low nanomolar to micromolar range in body fluids. Modification of the silica-wall with 1-(4-iodobutyl)4-aza-1-azoniabicyclo[2,2,2]octane iodide, also named M7C4I, has successfully been used to deactivate the fused silica wall and generate a stable reversed electroosmotic flow. Utilizing this advantage together with electrospray ionization time-of-flight mass spectrometry, which generates high resolution and fast acquisition monitoring of species, proved to be successful even for such a complex matrix like human cerebrospinal fluid.

  • 6.
    Barclay, Victoria K. H.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Tyrefors, Niklas L.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Johansson, I. Monika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt E.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Trace analysis of fluoxetine and its metabolite norfluoxetine: Part I: Development of a chiral liquid chromatography-tandem mass spectrometry method for wastewater samples2011In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1218, no 33, p. 5587-5596Article in journal (Refereed)
    Abstract [en]

    An enantioselective method for the determination of fluoxetine (a selective serotonin reuptake inhibitor) and its pharmacologically active metabolite norfluoxetine has been developed for raw and treated wastewater samples. The stable isotope-labeled fluoxetine and norfluoxetine were used in an extended way for extraction recovery calculations at trace level concentrations in wastewater. Wastewater samples were enriched by solid phase extraction (SPE) with Evolute CX-50 extraction cartridges. The obtained extraction recoveries ranged between 65 and 82% in raw and treated wastewater at a trace level concentration of 50 pM (15-16 ng L(-1)). The target compounds were identified by the use of chiral liquid chromatography tandem mass spectrometry (LC-MS/MS) in selected reaction monitoring (SRM) mode. The enantiomers were successfully resolved on a chiral alpha(1)-acid glycoprotein column (chiral AGP) with acetonitrile and 10 mM ammonium acetate buffer at pH 4.4 (3/97, v/v) as the mobile phase. The effects of pH, amount of organic modifier and buffer concentration in the mobile phase were investigated on the enantiomeric resolution (R(s)) of the target compounds. Enantiomeric R(s)-values above 2.0 (1.03 RSD%, n = 3) were achieved for the enantiomers of fluoxetine and norfluoxetine in all mobile phases investigated. The method was validated by assessing parameters such as cross-contamination and carryover during SPE and during LC analysis. Cross-talk effects were examined during the detection of the analytes in SRM mode. In addition, the isotopic purity of fluoxetine-d(5) and norfluoxetine-d(5) were assessed to exclude the possibility of self-contamination. The interassay precision of the chromatographic separation was excellent, with relative standard deviations (RSD) equal to or lower than 0.56 and 0.81% in raw and treated wastewaters, respectively. The method detection and quantification limits (respectively, MDL and MQL) were determined by the use of fluoxetine-d5 and norfluoxetine-d5. The MQL for the single enantiomers ranged from 12 to 14 pM (3.6-4.3 ng L(-1)) in raw wastewater and from 3 to 4 pM (0.9-1 ng L(-1)) in treated wastewater. The developed method has been employed for the quantification of (R)-fluoxetine, (S)-fluoxetine and the enantiomers of norfluoxetine in raw and treated wastewater samples to be presented in Part II of this study.

  • 7.
    Barclay, Victoria K H
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Tyrefors, Niklas L
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Johansson, I Monika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt E
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Trace analysis of fluoxetine and its metabolite norfluoxetine. Part II: Enantioselective quantification and studies of matrix effects in raw and treated wastewater by solid phase extraction and liquid chromatography-tandem mass spectrometry2012In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1227, p. 105-114Article in journal (Refereed)
    Abstract [en]

    The isotope-labeled compounds fluoxetine-d5 and norfluoxetine-d5 were used to study matrix effects caused by co-eluting compounds originating from raw and treated wastewater samples, collected in Uppsala, Sweden. The matrix effects were investigated by the determination of matrix factors (MF) and by a post-column infusion method. The matrix factors were determined to be 38–47% and 71–86% for the enantiomers of norfluoxetine-d5 and fluoxetine-d5, respectively. The influence of matrix effects when quantifying the enantiomers of the active pharmaceutical ingredient and the metabolite in wastewater samples with LC–MS/MS is discussed and methods to overcome the problem are presented. The enantiomeric concentrations of fluoxetine and its human metabolite norfluoxetine, quantified by a one-point calibration method, were 12–52 pM (3.5–16 ng L−1) in raw wastewater and 4–48 pM (1.2–15 ng L−1) in treated wastewater. Furthermore, the calculated enantiomeric fractions (EF) of the substances were found to be between 0.68 and 0.71 in both matrices. Neither the EF values for fluoxetine nor those for norfluoxetine were significantly different in the raw wastewater compared to the treated wastewater. Interestingly, the concentration of (S)-fluoxetine was found to be higher than the concentration of (R)-fluoxetine in both raw and treated wastewater. These results are different from other results presented in the literature, which shows that the relative concentrations of the enantiomers of a chiral active pharmaceutical ingredient might be significantly different in wastewater samples from different treatment systems. We report, for the first time, the concentrations of the enantiomers of norfluoxetine in wastewater samples. The concentrations of (S)-norfluoxetine were found to be higher than the concentration of (R)-norfluoxetine in the raw as well as in the treated wastewater samples.

  • 8.
    Barclay, Victoria K.H.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Tyrefors, Niklas L.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Johansson, I. Monika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Chiral analysis of metoprolol and two of its metabolites, alpha‑hydroxymetoprolol and deaminated metoprolol, in wastewater using liquid chromatography-tandem mass spectrometry2012In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1269, no SI, p. 208-217Article in journal (Refereed)
    Abstract [en]

    A LC–MS/MS method for the chiral separation of metoprolol and two of its main metabolites, α-hydroxymetoprolol (α-OH-Met) and deaminated metoprolol (COOH-Met), in environmental water samples has been developed. The target bases, metoprolol and α-OH-Met, as well as the acidic metabolite (COOH-Met) were extracted from water samples by a solid phase extraction method employing Oasis HLB cartridges. The extraction recoveries were ≥73% for all compounds in surface water. Four different types of chiral stationary phases were investigated for the separation of the eight stereoisomers of metoprolol and its metabolites, Chiralcel OD-H, Chirobiotic V, Chiral AGP and Chiral CBH. In the final method, the enantiomers of metoprolol and four stereoisomers of α-OH-Met were separated using Chiral CBH, the enantiomers of COOH-Met were separated employing Chiral AGP. The analytes were detected in SRM mode by triple quadrupole mass spectrometry. The method was applied for the chiral analysis of the analytes in treated wastewater samples from Uppsala, Sweden. The enantiomers and diastereoisomers of α-OH-Met were detected and analyzed in the samples. The concentrations of the three first eluting stereoisomers of α-OH-Met were between 54 and 61 pM. Interestingly, the last eluting stereoisomer was found to be present at a concentration of 151 pM at the same sampling occasion. This is, to the best of the authors’ knowledge, the first time the stereoisomers of α-OH-Met have been detected in wastewater samples. The enantiomers of metoprolol were determined to be 1.77 and 1.86 nM in the same matrix. The enantiomers of COOH-Met were not detected above the method detection limit (42 pM) in treated wastewater samples. The developed LC–MS/MS methods were validated in wastewater samples.

  • 9.
    Bergström, Sara K.
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Goiny, Michel
    Danielsson, Rolf
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Ungerstedt, Urban
    Andersson, Marit
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Markides, Karin E.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Screening of microdialysates taken before and after induced liver damage; on-line solid phase extraction-electrospray ionization-mass spectrometry2006In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1120, no 1-2, p. 21-26Article in journal (Refereed)
    Abstract [en]

    A novel method is described to follow known and unknown compounds in biological processes using microdialysis sampling and mass spectrometric detection. By implementation of internal standard, desalting/enrichment for the sample work-up, and multivariate data analysis, this methodology is a basis for future applications in early diagnosis of diseases and organ damage, as a complement to the routinely used clinical methods for biological samples. The present study includes screening without specific target analytes, of samples collected by microdialysis from liver of anaesthetized rats before and after local damage to this organ. Sample series were classified by principal component analysis, and the stimulation was identified in the chemical patterns produced by the presented analytical tool.

  • 10.
    Bäckström, Daniel
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Moberg, My
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Sjöberg, Per J. R.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Danielsson, Rolf
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Multivariate comparison between peptide mass fingerprints obtained by liquid chromatography-electrospray ionization-mass spectrometry with different trypsin digestion procedures2007In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1171, no 1-2, p. 69-79Article in journal (Refereed)
    Abstract [en]

    Peptide mass fingerprints were obtained for three different proteins using three different digestion procedures in triplicates with liquid chromatography coupled to electrospray ionization mass spectrometry. For each protein the results were compared with multivariate data analysis (cluster analysis, kernel principal component analysis) and pair-wise contrast evaluation. Clear systematic differences between the digestion procedures were established for all the proteins. The visual presentation of the pair-wise differences between procedures could to some extent be related to the protein fragments, although the main objective was to identify m/z and retention regions in the original peptide maps that should be subject to further exploration.

  • 11. Daszykowski, M.
    et al.
    Danielsson, Rolf
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Walczak, B.
    No-alignment-strategies for exploring a set of two-way data tables obtained from capillary electrophoresis-mass spectrometry2008In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1192, no 1, p. 157-165Article in journal (Refereed)
    Abstract [en]

    Hyphenated techniques such as capillary electrophoresis-mass spectrometry (CE-MS) or high-performance liquid chromatography with diode array detection (HPLC-DAD), etc., are known to produce a huge amount of data since each sample is characterized by a two-way data table. In this paper different ways of obtaining sample-related information from a set of such tables are discussed. Working with original data requires alignment techniques due to time shifts caused by unavoidable variations in separation conditions. Other pre-processing techniques have been suggested to facilitate comparison among samples without prior peak alignment, for example, 'binning' and/or 'blurring' the data along the time dimension. All these techniques, however, require optimization of some parameters, and in this paper an alternative parameter-free method is proposed. The individual data tables (X) are represented as Gram matrices (XXT), where the summation is taken over the time dimension. Hence the possible variations in time scale are eliminated, while the time information is at least partly preserved by the correlation structure between the detection channels. For comparison among samples, a similarity matrix is constructed and explored by principal component analysis and hierarchical clustering. The Gram matrix approach was tested and compared to some other methods using 'binned' and 'blurred' data for a data set with CE-MS runs on urine samples. In addition to data exploration by principal component analysis and hierarchical clustering, a discriminant partial least squares model was constructed to discriminate between the samples that were taken with and without the prior intake of a drug. The result showed that the proposed method is at least as good as the others with respect to cluster identification and class prediction. A distinct advantage is that there is no need for parameter optimization, while a potential drawback is the large size of the Gram matrices for data with high mass resolution.

  • 12.
    Edström, Lena
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Samuelsson, Jörgen
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Deformations of overloaded bands under pH-stable conditions in reversed phase chromatography2011In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1218, no 15, p. 1966-1973Article in journal (Refereed)
    Abstract [en]

    It has recently been demonstrated, using mathematical models, how peculiar overloaded band profiles of basic compounds are due to the local pH in the column when using low capacity buffers. In this study, overloaded peak shapes resulting after injection of carefully pH matched samples close to the pK(a) of the chosen solute are investigated primarily on two columns; one hybrid silica C18 column (Kromasil Eternity) and one purely polymeric column (PLRP-S), the latter lacking C18 ligands. It was found that distorted peaks of the basic test compound appear even though there is no difference in pH between the injected sample solution and the eluent; the previous explanation to why these effects occur is based on a pH mismatch. Thus, the unusual band shapes are not due to an initial pH difference. Furthermore, it was observed that the effect does not appear on polymeric columns without C18 ligands, but only on columns with C18 ligands, independently of the base matrix (silica, hybrid silica, polymeric).

  • 13.
    Elhamili, Anisa
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Wetterhall, Magnus
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Puerta, Angel
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Westerlund, Douglas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    The effect of sample salt additives on capillary electrophoresis analysis of intact proteins using surface modified capillaries.2009In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1216, no 17, p. 3613-3620Article in journal (Refereed)
    Abstract [en]

    The effect of adding alkali salts to protein samples for capillary electrophoretic (CE) analysis of intact proteins was studied. A high degree of peak stacking, even for large proteins, was found to occur when alkali salts were added to the sample. The addition of salt to the protein sample promotes a strong improvement in the peak efficiency of individual proteins giving up to 2.1 x 10(6) apparent plates/m. The concentration of salt required in the sample to reach optimal peak efficiency show dependency on both the molecular weight and molar concentration of the protein. However, adding salt will, at a sufficiently high concentration, cause a mixture of proteins to co-migrate to one very sharp peak. The observed sample stacking effect was obtained with a number of different surface modified silica capillaries indicating a general phenomenon and not surface coating specific.

  • 14.
    Elmongy, Hatem
    et al.
    Stockholm Univ, Dept Environm Sci & Analyt Chem, SE-10691 Stockholm, Sweden.;Damanhour Univ, Fac Pharm, Dept Pharmaceut Anal, Damanhour 22511, Egypt..
    Ahmed, Hytham
    Damanhour Univ, Fac Pharm, Dept Pharmaceut Anal, Damanhour 22511, Egypt..
    Wahbi, Abdel-Aziz
    Univ Alexandria, Fac Pharm, Dept Pharmaceut Analyt Chem, Alexandria 21521, Egypt..
    Koyi, Hirsh
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Centre for Research and Development, Gävleborg.
    Abdel-Rehim, Mohamed
    Stockholm Univ, Dept Environm Sci & Analyt Chem, SE-10691 Stockholm, Sweden..
    Online post-column solvent assisted and direct solvent-assisted electrospray ionization for chiral analysis of propranolol enantiomers in plasma samples2015In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1418, p. 110-118Article in journal (Refereed)
    Abstract [en]

    An Online post-column solvent-assisted ionization (OPSAI) method was developed for enhancing the ionization of the beta-blocker propranolol utilizing normal phase LC-MS/MS. Solvent-assisted electrospray ionization (SAESI) was studied by the introduction of the assistant solvents A: 0.5% Formic acid in Isopropanolol, B: 0.5% Formic acid in lsopropanolol-Water (1:1), and C: 0.5% Formic acid in water into the electrospray ionization chamber using a spray needle. Analyte molecules can be directly ionized by the aid of the assistant solvent spray. Both methods were applied to the chiral separation of propranolol enantiomers using normal phase analysis on cellulose-based chiral column. Interestingly, both methods are easy to handle and offer a wide range of assistant solvents that can be used in order to gain the optimum ionization of the analyte molecules. The both methods considerably improved the analyte signal and the peak area greatly increased. The propranolol average signal-to-noise (S/N) ratio was enhanced from 26 +/- 1 and 42 +/- 1 to 2341 +/- 61 and 1725 +/- 29 for R-propranolol and S-propranolol, respectively, when the post-column solvent method (OPSAI) was used with isopropanol-assistant solvent (A). While in case of solvent-assisted electrospray ionization method (SAESI) signal was enhanced from 26 +/- 1 and 42 +/- 1 to 2223 +/- 72 and 2155 +/- 58 for R-propranolol and S-propranolol, respectively, with water as an assistant solvent. The limit of detection was 10 ng/mL and the method was linear in the range 50-2000 ng/mL. The NPLC-MS method was applied for the determination of propranolol enantiomers in human plasma after microextraction by packed C18 sorbent.

  • 15.
    Enmark, Martin
    et al.
    Department of Chemistry and Biomedical Sciences, Karlstad University, Sweden.
    Samuelsson, Jörgen
    Department of Chemistry and Biomedical Sciences, Karlstad University, Sweden.
    Undin, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Characterization of an unusual adsorption behavior of racemic methyl-mandelate on a tris-(3,5-dimethylphenyl) carbamoyl cellulose chiral stationary phase2011In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1218, no 38, p. 6688-6696Article in journal (Refereed)
    Abstract [en]

    An interesting adsorption behavior of racemic methyl mandelate on a tris-(3,5-dimethylphenyl)carbamoyl cellulose chiral stationary phase was theoretically and experimentally investigated. The overloaded band of the more retained enantiomer had a peculiar shape indicating a type V adsorption isotherm whereas the overloaded band of the less retained enantiomer had a normal shape indicating a type I adsorption behavior. For a closer characterization of this separation, adsorption isotherms were determined and analyzed using an approach were Scatchard plots and adsorption energy distribution (AED) calculations are combined for a deeper analysis. It was found that the less retained enantiomer was best described by a Tóth adsorption isotherm while the second one was best described with a bi-Moreau adsorption isotherm. The latter model comprises non-ideal adsorbate–adsorbate interactions, providing an explanation to the non-ideal adsorption of the more retained enantiomer. Furthermore, the possibility of using the Moreau model as a local model for adsorption in AED calculations was evaluated using synthetically generated raw adsorption slope data. It was found that the AED accurately could predict the number of adsorption sites for the generated data. The adsorption behavior of both enantiomers was also studied at several different temperatures and found to be exothermic; i.e. the adsorbate–adsorbate interaction strength decreases with increasing temperature. Stochastic analysis of the adsorption process revealed that the average amount of adsorption/desorption events increases and the sojourn time decreases with increasing temperature.

  • 16.
    Eriksson, Anna
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bartsch, Maik
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Bergström Lind, Sara
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Agmo Hernández, Víctor
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    On-target titanium dioxide-based enrichment for characterization of phosphorylations in the Adenovirus pIIIa protein2013In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1317, no SI, p. 105-109Article in journal (Refereed)
    Abstract [en]

    A recently developed titanium dioxide (TiO2) based on-target method for phosphopeptide enrichment and matrix assisted laser desorption-ionization mass spectrometry (MALDI MS) analysis was used to investigate phosphorylations in the Adenovirus type 2 structural protein pIIIa. Lysates of purified virus particles were separated on 1-D SDS-PAGE and the band for the pIIIa protein was excised for tryptic digestion into peptides that were enriched with the on-target method. The enrichment provided by the method clearly improved the detectability of phosphorylated peptides and the results show for the first time evidence for multi-phosphorylated peptides in pIIIa. Moreover, three novel phosphorylations were identified in the protein sequence, even though the precise positions could not be determined. These results illustrate the potential of the method for the characterization of novel phosphoproteomes in biological samples of medical relevance.

  • 17.
    Fan, Ping
    et al.
    Department of Chemistry, University of Bremen AND School of Engineering and Science, Jacobs University Bremen, Germany.
    Neumann, Jennifer
    Center for Environmental Research and Sustainable Technology, University of Bremen, Germany.
    Stolte, Stefan
    Center for Environmental Research and Sustainable Technology, University of Bremen, Germany.
    Arning, Jürgen
    Center for Environmental Research and Sustainable Technology, University of Bremen, Germany.
    Ferreira, Denise
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC.
    Edwards, Katarina
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Gabel, Detlef
    Department of Chemistry, University of Bremen AND School of Engineering and Science, Jacobs University Bremen, Germany.
    Interaction of dodecaborate cluster compounds on hydrophilic column materials in water2012In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1256, p. 98-104Article in journal (Refereed)
    Abstract [en]

    The interaction of a series of dodecaborate cluster compounds B12X122− and B12X11Y2− (X = H, Cl, Br, I and Y = SH, OH, NR3) with hydrophilic column materials (Superdex 200, Sepharose 4B, Sephadex G-50, Sephadex G-100, alumina, silica gel and anion exchange material) was studied. Almost all the dodecaborate cluster compounds were retained strongly on Superdex 200. The halogenated cluster compounds interacted with Sepharose 4B, Sephadex G-50, Sephadex G-100 and alumina; on alumina, also the non-halogenated clusters were retained. Silica gel showed the least interaction with all compounds. The thermodynamic parameters were investigated for a selection of compounds on Superdex 200 and Sephadex G-100. Values for ΔH° were found to be negative on both gels. As the change in entropy ΔS° was also negative, it compensated ΔH° to a large extent. The clusters interacted also strongly with anion exchange material in ion chromatography; the interaction decreased with increasing acetonitrile concentration, implying a large contribution from solvent effects.

  • 18.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Characterization of adsorption processes in analytical liquid-solid chromatography2010In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1217, no 6, p. 792-812Article, review/survey (Refereed)
    Abstract [en]

    This review discusses nonlinear chromatographic methods of importance for proper characterization of the adsorption processes in analytical chromatographic systems, with focus on reversed-phase liquid chromatography. Linear methods such as the linear solvation energy relationship (LSER) method and the Snyder-Dolan hydrophobic-subtraction model will also be reviewed briefly. The nonlinear methods for adsorption isotherm determination and the tools for further treatment of the nonlinear adsorption data will be extensively treated in a way suitable for the general chromatographer. Applications of the various methods will be given and the outcome and conclusions will be discussed. Special emphasis will be placed on discussing the possibilities of combining linear and nonlinear methods in order to obtain a deeper and more complete investigation of the interactions in the actual phase system.

  • 19. Forssén, P.
    et al.
    Edström, Lena
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Lämmerhofer, M.
    Samuelsson, J.
    Karlsson, A.
    Lindner, W.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Analytical Chemistry.
    Optimization strategies accounting for the additive in preparative chiral liquid chromatography2012In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1269, no SI, p. 279-286Article in journal (Refereed)
    Abstract [en]

    This study is an in-depth investigation on how numerical optimization strategies that also account for the additive type and concentration, in preparative batch chromatography, should be performed. As a model system, the separation of Z-(R,S)-2-aminobutyric acid enantiomers on a quinidine carbamate-based chiral stationary phase in polar organic mode was used, with different additive strengths of acetic acid or hexanoic acid in methanol. The inverse method was used to determine the competitive adsorption isotherm parameters for the enantiomers and the additives. Three different optimization strategies were examined: (1) injection volume optimization, (2) optimization of injection volume and additive concentration, and (3) full optimization including injection volume, additive concentration, sample concentration and flow rate. It was concluded that (i) it is important to incorporate the additive concentration in the optimization procedure to achieve the highest production rates, (ii) the full optimization strategy had the overall best results, and (iii) the selection of additive is very important (here acetic acid additive was superior to the hexanoic acid additive). By including the additive in the adsorption model and in the numerical optimization it is not only possible to achieve higher production rates but also to properly select the additive that is most advantageous for the specific separation problem.

  • 20.
    Forssén, Patrik
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Arnell, R.
    Kaspereit, M.
    Seidel-Morgenstern, A.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Effects of a strongly adsorbed additive on process performance in chiral preparative chromatography2008In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1212, no 1-2, p. 89-97Article in journal (Refereed)
    Abstract [en]

    The shapes of elution profiles are often significantly influenced by the presence of strongly adsorbed additives in the mobile phase. This aspect needs to be considered in quantitative optimization of preparative chromatography. The theoretical study carried out here is based on available thermodynamic information for the enantiomers of three beta-blockers, alprenolol, propranolol, and atenolol, on a teicoplanin chiral stationary phase (Chirobiotic T) using methanol/acetonitrile as the mobile phase and acetic acid/triethylamine as the additive. The properties of this strong additive made it possible to tune the binary elution profiles in any combination of the following apparent band shapes: anti-Langmuir/anti-Langmuir, anti-Langmuir/Langmuir and Langmuir/Langmuir. Optimization of the productivity and yield, when performing repetitive batch injections, was investigated using the equilibrium dispersive model. We show that it is important to consider the invisible additive perturbation peak when defining the cycle time and therefore a model-based optimization needs to take this into account. Furthermore, both productivity and yield could be improved for the two unusual shape combinations in comparison to the traditional Langmuir/Langmuir case.

  • 21.
    Forssén, Patrik
    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.
    A quest for the optimal additive in chiral preparative chromatography2009In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1216, no 23, p. 4719-4727Article in journal (Refereed)
    Abstract [en]

    Traditionally, the choice of acid/base additives used in chiral preparative chromatography has not been considered very important. However, it was recently demonstrate that strongly adsorbing additives can result in the Most unexpected enantiomer band shapes in modern chiral preparative chromatographic systems. In the present study we demonstrate that, depending on the choice of additive, it is actually possible to obtain the following four binary band-shape compositions when a racemic mixture is injected: (i) anti-Langmuir/anti-Langmuir, (ii) anti-Langmuir/Langmuir, (iii) Langmuir/Langmuir and (iv) Langmuir/anti-Langmuir. Further, we made an advanced numerical investigation, in order to ascertain which one of the four band-shape compositions, is the most favourable one in preparative batch chromatography of a racemic mixture. We found that if the target for purification is either the first eluting enantiomer or both ones, the traditional Langmuir/Langmuir band-shape composition should be chosen. But, if only the second eluting enantiomer is to be purified the optimal situation is the anti-Langmuir/Langmuir band-shape composition. Thus, it was concluded that the best choice of additive depends on which enantiomer is of interest and it is useful to perform a thorough additive screening to find the optimal additive, giving the most advantageous peak shape composition and accordingly the best process performance for a particular separation problem. (C) 2009 Elsevier B.V. All rights reserved.

  • 22.
    Forssén, Patrik
    et al.
    Department of Chemistry and Biomedical Sciences, Karlstad University, Sweden.
    Edström, Lena
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Samuelsson, Jörgen
    Department of Chemistry and Biomedical Sciences, Karlstad University, Sweden.
    Fornstedt, Torgny
    Department of Chemistry and Biomedical Sciences, Karlstad University, Sweden.
    Injection profiles in liquid chromatography II: predicting accurate injection-profiles for computer-assisted preparative optimizations.2011In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1218, no 34, p. 5794-5800Article in journal (Refereed)
    Abstract [en]

    In computer assisted optimization of liquid chromatography it has been known for some years that it is important to use experimental injection profiles, instead of rectangular ones, in order to calculate accurate elution bands. However, the incorrectly assumed rectangular profiles are still mostly used especially in numerical optimizations. The reason is that the acquisition of injection profiles, for each injection volume and each flow rate considered in a computer-assisted optimization requires a too large number of experiments. In this article a new function is proposed, which enables highly accurate predictions of the injection profiles and thus more accurate computer optimizations, with a minimum experimental effort. To model the injection profiles for any injection volume at a constant flow rate, as few as two experimental injection profiles are required. If it is desirable to also take the effect of flow rate on the injection profiles into account, then just two additional experiments are required. The overlap between fitted and experimental injection profiles at different flow rates and different injection volumes were excellent, more than 90%, using experimental injection profiles from just four different injection volumes at two different flow rates. Moreover, it was demonstrated that the flow rate has a minor influence on the injection profiles and that the injection volume is the main parameter that needs to be accounted for.

  • 23.
    Forssén, Patrik
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    General theory of indirect detection in chromatography2006In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1126, p. 268-275Article in journal (Refereed)
  • 24.
    Forssén, Patrik
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Division of Scientific Computing. Uppsala University, Disciplinary Domain of Science and Technology, Mathematics and Computer Science, Department of Information Technology, Numerical Analysis.
    Lindholm, Johan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology.
    Theoretical and experimental study of binary perturbation peaks with focus on peculiar retention behaviour and vanishing peaks in chiral liquid chromatography2003In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 991, p. 31-45Article in journal (Refereed)
  • 25. Franco Fraguas, L
    et al.
    Carlsson, Jan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Lönnberg, Maria
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Lectin affinity chromatography as a tool to differentiate endogenous and recombinant erythropoietins2008In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1212, no 1-2, p. 82-88Article in journal (Refereed)
    Abstract [en]

    This work exploits the combination of the lectin affinity chromatography (LAC) with an ultra-sensitive immunochromatographic assay to differentiate several types of erythropoietin (EPO). The chromatographic behaviours of different commercial types of recombinant human EPO (rhEPO), EPO analogues (Aranesp) and urine human EPO (uhEPO) from healthy individuals on eight lectin-Sepharose columns, have been worked out. Results show that when using wheat germ agglutinin (WGA)-Sepharose columns, a careful desorption regime starting with very low concentration (2mM) of the competitive sugar N-acetylglucosamine (GlcNAc) makes it possible to efficiently distinguish endogenous EPO from recombinant EPO and EPO analogues.

  • 26. Gonzalez-Ortega, Omar
    et al.
    Porath, Jerker
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine.
    Guzman, Roberto
    Adsorption of peptides and small proteins with control access polymer permeation to affinity binding sites. Part I: Polymer permeation-immobilized metal ion affinity chromatography separation adsorbents with polyethylene glycol and immobilized metal ions2012In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1227, p. 115-125Article in journal (Refereed)
    Abstract [en]

    Despite the many efforts to develop efficient protein purification techniques, the isolation of peptides and small proteins on a larger than analytical scale remains a significant challenge. Recovery of small biomolecules from diluted complex biological mixtures, such as human serum, employing porous adsorbents is a difficult task mainly due to the presence of concentrated large biomolecules that can add undesired effects in the system such as blocking of adsorbent pores, impairing diffusion of small molecules, or competition for adsorption sites. Adsorption and size exclusion chromatography (AdSEC) controlled access media, using polyethylene glycol (PEG) as a semi-permeable barrier on a polysaccharide matrix, have been developed and explored in this work to overcome such effects and to preferentially adsorb small molecules while rejecting large ones. In the first part of this work, adsorption studies were performed with small peptides and proteins from synthetic mixtures using controlled access polymer permeation adsorption (CAPPA) media created by effectively grafting PEG on an immobilized metal affinity chromatography (IMAC) agarose resin, where chelating agents and immobilized metal ions were used as the primary affinity binding sites. Synthetic mixtures consisted of bovine serum albumin (BSA) with small proteins, peptides, amino acids (such as histidine or Val(4)-Angiotensin III), and small molecules-spiked human serum. The synthesized hybrid adsorbent consisted of agarose beads modified with iminodiacetic (IDA) groups, loaded with immobilized Cu(II) ions, and PEG. These CAPPA media with grafted PEG on the interior and exterior surfaces of the agarose matrix were effective in rejecting high molecular weight proteins. Different PEG grafting densities and PEG of different molecular weight were tested to determine their effect in rejecting and controlling adsorbent permeation properties. Low grafting density of high molecular weight PEG was found to be as effective as high grafting density of low molecular weight PEG in the rejecting properties of the semi-permeable synthesized media. (C) 2012 Elsevier B.V. All rights reserved.

  • 27. Gonzalez-Ortega, Omar
    et al.
    Porath, Jerker
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine.
    Guzman, Roberto
    Adsorption of peptides and small proteins with control access polymer permeation to affinity binding sites. Part II: Polymer permeation-ion exchange separation adsorbents with polyethylene glycol and strong anion exchange groups2012In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1227, p. 126-137Article in journal (Refereed)
    Abstract [en]

    In chromatographic separations, the most general problem in small biomolecule isolation and purification is that such biomolecules are usually found in extremely low concentrations together with high concentrations of large molecular weight proteins. In the first part of this work, adsorption and size exclusion chromatography (AdSEC) controlled access media, using polyethylene glycol (PEG) as a semi-permeable barrier on a polysaccharide Immobilized Metal Affinity Chromatography (IMAC) matrix was synthesized and used to develop chromatographic adsorbents that preferentially adsorb and separate low molecular weight biomolecules while rejecting large molecular weight proteins. In this second part, we expand the concept of controlled access polymer permeation adsorption (CAPPA) media by grafting polyethylene glycol (PEG) on a high capacity polysaccharide ion exchange (IEX) chromatographic resin where PEG acts as a semi-permeable barrier that preferentially allows the permeation of small molecules while rejecting large ones. The IEX resin bearing quaternary ammonium groups binds permeated biomolecules according to their ion exchange affinity while excluding large biomolecules by the PEG barrier and thus cannot compete for the binding sites. This new AdSEC media was used to study the retention of peptides and proteins covering a wide range of molecular weights from to 150 kDa. The effect of protein molecular weight towards retention by ion exchange was performed using pure protein solutions. Recovery of insulin from insulin-spiked human serum and insulin-spiked human urine was evaluated under polymer controlled permeation conditions. The CAPPA media consisted of agarose beads modified with amino-PEG-methoxy and with trimethyl ammonium groups, having chloride capacities between 20 and 40 mu eq/mL and were effective in rejecting high molecular weight proteins while allowing the preferential adsorption of small proteins and peptides. 

  • 28.
    Gyllenhaal, Olle
    et al.
    Analytical & Technical Development, Pharmaceutical and Analytical R&D, AstraZeneca R&D Mölndal, Sweden.
    Edström, Lena
    Analytical & Technical Development, Pharmaceutical and Analytical R&D, AstraZeneca R&D Mölndal, Sweden.
    Persson, Bengt-Arne
    Development DMPK & Bioanalysis Mölndal, AstraZeneca R&D Mölndal, Sweden.
    Ion-pair supercritical fluid chromatography of metoprololand related amino alcohols on diol silica2006In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1134, p. 305-310Article in journal (Refereed)
    Abstract [en]

    In this paper, a chromatographic system based on carbon dioxide with methanol as mobile phase, and diol silica as stationary phase has been investigated for metoprolol and related amino alcohols by addition of strong acids to systems with triethylamine base as primary additive. Standard conditions used were 10% of methanol, containing 24mM of acid and 18mM of triethylamine, in carbon dioxide with a flow rate of 1.5 ml min−1. The column dimensions were 125mm×4mm I.D. and kept at 40 ◦C with a back pressure of 150 bar. Effects on selectivity were stronger with trifluoroacetic acid than with ethanesulfonic acid. From a large set of related analytes, it was shown that selectivity changes were significant when the structure close to the nitrogen of the amino alcohol analyte differed. The stability of the column in the short time perspective was examined and it showed negligible changes. For a diastereoisomeric pair, not resolved in a basic system with triethylamine nor by addition of ethanesulfonic acid, resolution improved to about 2.1 with trifluoroacetic acid. The described approach offers a way to tune the selectivity of SFC systems when amines are analyzed without the need to change stationary phase for the chromatographic separation.

  • 29.
    Götmar, Gustaf
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Samuelsson, Jörgen
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Karlsson, Anders
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Thermodynamic characterization of the adsorption of selected chiral compounds on immobilized amyloglucosidase in liquid chromatography2007In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1156, no 1-2, p. 3-13Article in journal (Refereed)
    Abstract [en]

    Immobilized amyloglucosidase was used as a chiral stationary phase (CSP). First, the retention and enantioselectivity of several model chiral amines and acids were investigated. We found that this CSP was unable to separate the enantiomers of acids, though all selected amines could be resolved. The adsorption of (R)- and (S)-propranolol and its influence on column temperature and 2-propanol content in the eluent were then studied in detail, using a three-step methodology. The adsorption was first evaluated using Scatchard plots; thereafter, the adsorption was characterized in detail by calculating the adsorption energy distribution. With this model-independent information, a better judgment could be made of the possible adsorption models selected in the last step, the model fitting to the data. In the case examined, the bi-Langmuir model (containing nonselective and enantioselective sites) describes the system well. The retention of (R)- and (S)-propranolol at low temperatures increases with the content of 2-propanol in the eluent, due to the increased saturation capacity of the enantioselective sites. The retention is an enthalpy-driven process at both types of sites, whereas the enantioseparation is due to differences between the entropy changes of the two enantiomers at the enantioselective sites. The enthalpy of adsorption at the nonselective sites is almost identical at the two concentrations of 2-propanol in the eluent. Enantioselective adsorption, on the other hand, is more exothermic at higher modifier content (20%). Thus, at high temperatures the retention decreases with increasing modifier content, whereas the opposite (unusual) trend is the case at low temperatures.

  • 30.
    Han, Xiao
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Pathmasiri, Wimal
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Bohlin, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Janson, Jan-Christer
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Isolation of high purity 1-[2′,4′-dihydroxy-3′,5′-di-(3″-methylbut-2″-enyl)-6′-methoxy] phenylethanone from Acronychia pedunculata (L.) Miq. by high-speed counter-current chromatography2004In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1022, no 1-2, p. 213-216Article in journal (Refereed)
    Abstract [en]

    Following an initial clean-up step on silica, high-speed counter-current chromatography (HSCCC) was used to purify an aryl ketone, 1-[2′,4′-dihydroxy-3′,5′-di-(3″-methylbut-2″-enyl)-6′-methoxy] phenylethanone from an extract of the stem bark of the shrub Acronychia pedunculata. The two-phase solvent system used was composed of n-heptane–ethyl acetate–methanol–water at an optimized volume ratio of 4:1:4:1 (v/v/v/v). Target compound (58.1 mg) with a purity of 98.9% was obtained after HSCCC of 183.5 mg sample with a purity of 35.7% recovered after the silica clean-up step. Identification of the target compound was performed by 1H NMR, 13C NMR, two-dimensional NMR and LC–electrospray ionization MS.

  • 31.
    Hanrieder, Jörg
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Zuberovic, Aida
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Bergquist, Jonas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Surface modified capillary electrophoresis combined with in solution isoelectric focusing and MALDI-TOF/TOF MS: A gel-free multidimensional electrophoresis approach for proteomic profiling — Exemplified on human follicular fluid2009In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1216, no 17, p. 3621-3628Article in journal (Refereed)
    Abstract [en]

    Development of miniaturized analytical tools continues to be of great interest to face the challenges in proteomic analysis of complex biological samples such as human body fluids. In the light of these challenges, special emphasis is put on the speed and simplicity of newly designed technological approaches as well as the need for cost efficiency and low sample consumption. In this study, we present an alternative multidimensional bottom-up approach for proteomic profiling for fast, efficient and sensitive protein analysis in complex biological matrices. The presented setup was based on sample pre-fractionation using microscale in solution isoelectric focusing (IEF) followed by tryptic digestion and subsequent capillary electrophoresis (CE) coupled off-line to matrix assisted laser desorption/ionization time of flight tandem mass spectrometry (MALDI TOF MS/MS). For high performance CE-separation, PolyE-323 modified capillaries were applied to minimize analyte-wall interactions. The potential of the analytical setup was demonstrated on human follicular fluid (hFF) representing a typical complex human body fluid with clinical implication. The obtained results show significant identification of 73 unique proteins (identified at 95% significance level), including mostly acute phase proteins but also protein identities that are well known to be extensively involved in follicular development.

  • 32.
    Hedeland, Ylva
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Hedeland, Mikael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Bondesson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Chiral separation of amines with N-benzoxycarbonylglycyl-L-proline as selector in non-aqueous capillary electrophoresis using methanol and 1,2-dichloroethane in the background electrolyte2003In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 984, no 2, p. 261-271Article in journal (Refereed)
    Abstract [en]

    N-Benzoxycarbonylglycyl-L-proline (L-ZGP) has been introduced as a chiral selector for enantioseparation of amines in non-aqueous capillary electrophoresis. Methanol mixed with different proportions of dichloromethane, 1,2-dichloroethane or 2-propanol containing L-ZGP and ammonium acetate was used as the background electrolyte. Enantioseparation of different types of pharmacologically active amines was performed, e.g. the local anaesthetic bupivacaine and the beta-adrenoceptor blocking agent pindolol. Addition of the solvents (dichloromethane, 1,2-dichloroethane or 2-propanol) gave an improved chiral separation partly due to a distinct decrease in the electroosmotic flow. The use of 1,2-dichloroethane in the background electrolyte gave higher precision in migration time (RSD 2.2%) compared to the systems containing dichloromethane. An enantiomeric separation of mepivacaine was performed within 72 s by use of short-end injection with an effective capillary length of 8.5 cm.

  • 33.
    Hedeland, Ylva
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Lehtinen, Jenni
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Ketopinic acid and diisoproylideneketogulonic acid as chiral ion-pair selectors in capillary electrophoresis: Enantiomeric impurity analysis of S-timolol and 1R,2S-ephedrine2007In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1141, no 2, p. 287-294Article in journal (Refereed)
    Abstract [en]

    1S,4R-(+)-ketopinic acid [(+)-KPA] has been introduced as a chiral selector for the separation of pharmacologically active amines by non-aqueous capillary electrophoresis (NACE). (+)-KPA gave enantioresolution for most of the compounds previously separated by 2R,3S,4R,5S-(−)-2,3:4,6-di-O-isopropylidene-2-keto-l-gulonic acid [(−)-DIKGA], but with a reversed migration order. A complete enantioresolution (Rs = 4.2) was obtained for timolol, a compound that could not be resolved using (−)-DIKGA as the selector. Thus, (+)-KPA was evaluated for the enantiomeric purity determination of S-timolol. A method based on pre-concentration by transient isotachophoresis (tITP) provided a limit of detection (LOD) of 0.2% R-timolol in S-timolol samples. Because of the lack of enantioresolution of ephedrine when (+)-KPA was used as the selector, a method with (−)-DIKGA has been developed and validated for determination of the enantiomeric purity of the 1R,2S enantiomer. The method gave good precision and accuracy with an LOD (S/N = 3) of 0.033% for the enantiomeric impurity 1S,2R-ephedrine.

  • 34.
    Hedeland, Ylva
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Lethinen, Jenni
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Determination of the enantiomeric impurity in 1R,2S-ephedrine and S-timolol by chiral ion-pair non-aqueous capillary electrophoresis2007In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1141, p. 287-294Article in journal (Refereed)
    Abstract [en]

    1S,4R-(+)-ketopinic acid [(+)-KPA] has been introduced as a chiral selector for the separation of pharmacologically active amines by non-aqueouscapillary electrophoresis (NACE). (+)-KPA gave enantioresolution for most of the compounds previously separated by 2R,3S,4R,5S-(−)-2,3:4,6-di-O-isopropylidene-2-keto-l-gulonic acid [(−)-DIKGA], but with a reversed migration order. A complete enantioresolution (Rs = 4.2) was obtainedfor timolol, a compound that could not be resolved using (−)-DIKGA as the selector. Thus, (+)-KPA was evaluated for the enantiomeric puritydetermination of S-timolol. A method based on pre-concentration by transient isotachophoresis (tITP) provided a limit of detection (LOD) of0.2% R-timolol in S-timolol samples. Because of the lack of enantioresolution of ephedrine when (+)-KPA was used as the selector, a methodwith (−)-DIKGA has been developed and validated for determination of the enantiomeric purity of the 1R,2S enantiomer. The method gave goodprecision and accuracy with an LOD (S/N = 3) of 0.033% for the enantiomeric impurity 1S,2R-ephedrine.

  • 35.
    Henriksson, Hongbin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Munoz, Irene
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Molecular Biology.
    Isaksson, Roland
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Chemistry.
    Pettersson, Göran
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Johansson, Gunnar
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Cellobiohydrolase 58 (P.c. Cel 7D) is complementary to the homologous CBHI (T.r. Cel 7A) in enantioseparations2000In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 898, no 1, p. 63-74Article in journal (Refereed)
    Abstract [en]

    Cellobiohydrolase 58 (EC 3.2.1.91, pc. Cel 7D) from Phanerochaete chrysosporium was immobilized on silica and the resulting material, CBH 58-silica, was then used as a chiral stationary phase (CSP) in liquid chromatographic separations of enantiomers. The enantioselectivities obtained on CBH 58-silica were compared with those on CBH I-silica (a phase based on a corresponding cellulase from Trichoderma reesei). CBH 58-silica displayed higher selectivity than CBH I-silica for the more hydrophilic compounds, such as atenolol and metoprolol, although great similarities in chiral separation of beta -adrenergic antagonists were found between the two phases. None of the acidic compounds tested could be resolved on the CBH 58 phase. Moreover, the solutes were retained more on the CBH 58 phase in general, indicating an improved application potential in bioanalysis. Addition of cellobiose or lactose, both of which are inhibitors of cellulases, To the mobile phase impaired the enantioselectivity, indicating an overlap of the enantioselective and catalytic sites. The chiral analytes also functioned as competitive inhibitors and their inhibition constants were determined.

  • 36.
    Henriksson, Hongbin
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC.
    Pettersson, Göran
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC.
    Johansson, Gunnar
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
    Discrimination between enantioselective and non-selective binding sites on cellobiohydrolase-based stationary phases by selective displacers1999In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 857, p. 107-115Article in journal (Refereed)
  • 37. Iadaresta, Francesco
    et al.
    Crescenzi, Carlo
    Amini, Ahmad
    Colmsjö, Anders
    Koyi, Hirsh
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Centre for Research and Development, Gävleborg.
    Abdel-Rehim, Mohamed
    Application of graphitic sorbent for online microextraction of drugs in human plasma samples2015In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1422, p. 34-42Article in journal (Refereed)
    Abstract [en]

    In the present work a new sorbent based on graphitized carbon (CarbonX(®) COA) was evaluated in microextraction by packed sorbent (MEPS) for extraction of lidocaine and ropivacaine from human plasma samples. The new graphitic sorbent showed high recoveries of lidocaine and ropivacaine compared to C18 sorbent. In the present study the G-MEPS (syringe packed with graphitic sorbent) was connect online with liquid chromatography tandem mass spectrometry (LC-MS/MS). In order to obtain a fast and reliable method different factors affecting MEPS performance were investigated. The extraction efficiency of the graphitic sorbent was compared with silica-based sorbents used in MEPS. The G-MEPS was also evaluated for reuse (50-100 times). The recoveries of lidocaine and ropivacaine from plasma samples were 79% and 82%; respectively. The method was validated according to FDA (Food and Drug Administration) guideline for bioanalytical method validation. Linearity was assessed in the range 5-2000nmol/L, with coefficient of determination r(2)>0,995 (n=3) for lidocaine and r(2)>0.997 (n=3) for ropivacaine. The lower limit of quantification (LLOQ) was 5nmol/L and the limit of detection (LOD) was 1nmol/L for studied analytes in plasma samples. For both analytes considered in this study the accuracy values in plasma samples were ranged from 86% to 113%. The Inter-day precisions, expressed as relative standard deviation (%RSD), at three different concentrations (QC-samples) ranged from 8% to 9% for lidocaine, and from 4% to 11% for ropivacaine.

  • 38. Jansson, Christer
    et al.
    Pihlström, Tuija
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Analytical Chemistry.
    Österdahl, Bengt-Göran
    Markides, Karin E
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Analytical Chemistry.
    A new multi-residue method for analysis of pesticide residues in fruit and vegetables using liquid chromatography with tandem mass spectrometric detection2004In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1023, no 1, p. 93-104Article in journal (Refereed)
    Abstract [en]

    A new multi-residue method for determination of pesticide residues in a wide variety of fruit and vegetables, using the National Food Administration (NFA) ethyl acetate extraction and determination by means of LC-MS/MS, is presented. The method includes pesticides normally detected by LC-UV or LC-fluorescence such as benzimidazoles, carbamates, N-methylcarbamates and organophosphorus compounds with an oxidisable sulphide group as well. After extraction with ethyl acetate, the extract is concentrated and an aliquot of the extract is evaporated to dryness and redissolved in methanol before injection on LC-MS/MS. The method has been validated for 57 different pesticides and metabolites. Representative species from different commodity groups were chosen as matrices in order to study the influence from different matrices on recoveries. The fortification levels studied were 0.01-0.5 mg kg(-1). Matrix effects were tested for all matrices by means of standard addition to blank extracts. The matrix effect, expressed as signal in solvent compared to signal in matrix, was in general found to be small. The obtained recoveries are, with a few exceptions, in the range 70-100%. The proposed method is quick and straightforward and no additional clean-up steps are needed. The method can be used for the analysis of all 57 pesticides in one single determination step at 0.01 mg kg(-1).

  • 39. Johansson, Bo-Lennart
    et al.
    Andersson, Mikael
    Lausmaa, Jukka
    Sjövall, Peter
    Chemical characterisation of different separation media based on agarose by static time-of-flight secondary ion mass spectrometry2004In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1023, no 1, p. 49-56Article in journal (Refereed)
    Abstract [en]

    In this paper, the novel application of time-of-flight secondary ion mass spectrometry (TOF-SIMS) for qualitative and semi-quantitative investigation of the surface chemistry of separation media based on beaded agarose is reported. Five different media were studied: DEAE Sepharose Fast Flow, Q Sepharose Fast Flow, SP Sepharose Fast Flow, Phenyl Sepharose Fast Flow at ligand densities between 7 and 33% (w/w) and the base matrix Sepharose 6 Fast Flow. The obtained TOF-SIMS spectra reveal significant chemical information regarding the ligands (DEAE, Q, SP and Phenyl) which are covalently attached to the agarose-based matrix Sepharose 6 Fast Flow. For the anion-exchange media (DEAE and Q Sepharose Fast Flow), the positive TOF-SIMS spectra yielded several strong characteristic fragment peaks from the amine ligands. Structural information was obtained, e.g. from the peak at m/z 173.20, originating from the ion structure [(C2H5)2NCH2CH2NH(C2H5)2l+, which shows that the ligand in DEAE Sepharose Fast Flow is composed of both tertiary and quaternary amines. The positive spectrum of Phenyl Sepharose Fast Flow contained major fragments both from the base matrix and the ligand. The cation-exchanger (SP Sepharose Fast Flow) gave rise to a positive spectrum resembling that of the base matrix (Sepharose 6 Fast Flow) but with a different intensity pattern of the matrix fragments. In addition, peaks with low intensity at m/z 109.94, 125.94 and 139.95 corresponding to Na2SO2+, Na2SO3+ and Na2SO3CH2+, respectively, were observed. The positive TOF-SIMS spectrum of Sepharose 6 Fast Flow contains a large number of fragments in the mass range up to m/z 200 identified as CxHyOz and CxHy structures. The results clearly show that positive TOF-SIMS spectra of different media based on Sepharose 6 Fast Flow are strongly influenced by the ligand coupled to the matrix. The negative TOF-SIMS spectra contained several ligand-specific, characteristic peaks for the cation-exchanger, having sulphonate as the ion-exchange group. Negative fragments such as S-, SO-, SO2-, SO3-, C2H3SO3-, C3H5SO3- and OC3H5SO3- were observed. Phenyl Sepharose Fast Flow, which has an uncharged group (Phenyl) coupled to the agarose matrix yielded one ligand-related peak corresponding to the C6H5O- fragment. DEAE and Q ligands could only be identified by the appearance of the fragments CN- and CNO- in the negative spectrum. However, a strong peak corresponding to the counter ion (Cl-) was observed. TOF-SIMS analysis can also be used for the investigation of residues from the coupling procedure that bonds the ligands to the matrix. One example is the observation of bromine peaks in the negative spectrum of Q Sepharose Fast Flow. Furthermore, it has also been shown that different ligand concentrations of Phenyl Sepharose Fast Flow can easily be detected by TOF-SIMS analysis. Information regarding the difference between the ligand density on the surface of the beads and in the bulk can also be obtained. However, spectra registered on the outermost surface and on the pore surface (crushed beads) of DEAE Sepharose Fast Flow clearly show that the agarose and the DEAE groups are homogeneously distributed in the beads.

  • 40.
    Lindholm, Johan
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Westerlund, Douglas
    Karlsson, Karl-Erik
    Caldwell, Karin
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology.
    Fornstedt, Torgny
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Surface Biotechnology. Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry.
    Use of liquid chromatography-diode-array detection and mass spectrometry for rapid product identification in biotechnological synthesis of a hydroxyprogesterone2003In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 992, no 1-2, p. 85-100Article in journal (Refereed)
    Abstract [en]

    In exploratory scale biotechnological process development, the product must be rapidly identified although a reference compound may not always be available. LC-diode-array detection and MS were used for this purpose in a process producing 9alpha-hydroxyprogesterone from progesterone as substrate. The electrospray ionization mass spectrometer was combined with an ion trap mass spectrometer for the second generation MS. The preliminary identification, which could be carried out within the course of a day, confirmed that the product was a hydroxyprogesterone. The final identification step, which was much more material intensive and hence time consuming, involved a two-step preparative separation to yield quantities necessary for definitive product identification based on 1H- and 13C NMR.

  • 41.
    Liu, Xiaojie
    et al.
    Leiden Univ, Inst Biol, Nat Prod Lab, NL-2333 BE Leiden, Netherlands..
    Ahlgren, Samantha
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Farmakognosi. Leiden Univ, Inst Biol, Nat Prod Lab, NL-2333 BE Leiden, Netherlands.
    Korthout, Henrie A. A. J.
    Fytagoras BV, NL-2333 BE Leiden, Netherlands..
    Salome-Abarca, Luis F.
    Leiden Univ, Inst Biol, Nat Prod Lab, NL-2333 BE Leiden, Netherlands..
    Bayona, Lina M.
    Leiden Univ, Inst Biol, Nat Prod Lab, NL-2333 BE Leiden, Netherlands..
    Verpoorte, Robert
    Leiden Univ, Inst Biol, Nat Prod Lab, NL-2333 BE Leiden, Netherlands..
    Choi, Young Hae
    Leiden Univ, Inst Biol, Nat Prod Lab, NL-2333 BE Leiden, Netherlands.;Kyung Hee Univ, Coll Pharm, Seoul 02447, South Korea..
    Broad range chemical profiling of natural deep eutectic solvent extracts using a high performance thin layer chromatography-based method2018In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1532, p. 198-207Article in journal (Refereed)
    Abstract [en]

    Natural deep eutectic solvents (NADES) made mainly with abundant primary metabolites are being increasingly applied in green chemistry. The advantages of NADES as green solvents have led to their use in novel green products for the food, cosmetics and pharma markets. However, one of the main difficulties encountered in the development of novel products and their quality control arises from their low vapour pressure and high viscosity. These features create the need for the development of new analytical methods suited to this type of sample. In this study, such a method was developed and applied to analyse the efficiency of a diverse set of NADES for the extraction of compounds of interest from two model plants, Ginkgo biloba and Panax ginseng. The method uses high-performance thin-layer chromatography (HPTLC) coupled with multivariate data analysis (MVDA). It was successfully applied to the comparative quali- and quantitative analysis of very chemically diverse metabolites (e.g., phenolics, terpenoids, phenolic acids and saponins) that are present in the extracts obtained from the plants using six different NADES. The composition of each NADES was a combination of two or three compounds mixed in defined molar ratios; malic acid-choline chloride (1:1), malic acid-glucose (1:1), choline chloride-glucose (5:2), malic acid-proline (1:1), glucose-fructose-sucrose (1:1:1) and glycerol-proline-sucrose (9:4:1). Of these mixtures, malic acid-choline chloride (1:1) and glycerol-proline-sucrose (1:1:1) for G. biloba leaves, and malic acid-choline chloride (1:1) and malic acid-glucose (1:1) for P. ginseng leaves and stems showed the highest yields of the target compounds. Interestingly, none of the NADES extracted ginkgolic acids as much as the conventional organic solvents. As these compounds are considered to be toxic, the fact that these NADES produce virtually ginkgolic acid-free extracts is extremely useful. The effect of adding different volumes of water to the most efficient NADES was also evaluated and the results revealed that there is a great influence exerted by the water content, with maximum yields of ginkgolides, phenolics and ginsenosides being obtained with approximately 20% water (w/w).

  • 42.
    Lodén, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Hedeland, Ylva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Hedeland, Mikael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Bondesson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Development of a chiral non-aqueous capillary electrophoretic system using the partial filling technique with UV and mass spectrometric detection2003In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 986, no 1, p. 143-152Article in journal (Refereed)
    Abstract [en]

    A chiral non-aqueous CE system with UV and mass spectrometric detection has been developed. The enantioseparation was promoted by diastereomeric complex (ion-pair) formation between the amines (e.g. salbutamol, atenolol) and the chiral selector, (-)-2,3:4,6-di-O-isopropylidene-2-keto-L-gulonic acid [(-)-DIKGA]. Different solvent mixtures were studied, as well as different concentrations of (-)-DIKGA and ammonium acetate in the background electrolyte. A partial filling technique was developed with a selector plug composed of (-)-DIKGA and ammonium acetate in a solvent mixture of methanol and 2-propanol. The separated enantiomers of pronethalol were detected by a Q-TOF MS system equipped with a sheath-flow electrospray ionization interface.

  • 43.
    Lodén, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Arvidsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Amini, Ahmad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Quantitative determination of salbutamol in tablets by multiple-injection capillary zone electrophoresis2008In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1207, no 1-2, p. 181-185Article in journal (Refereed)
    Abstract [en]

    A multiple-injection capillary zone electrophoresis (MICZE) method has been developed for the assay of salbutamol in Ventoline Depot tablets (GlaxoSmithKline). In the developed method, seven sample sets, each consisting of three samples, were sequentially injected into the capillary and analyzed within a single run. This enabled a total of twenty-one sequential injections, i.e., six standards and fifteen samples, containing salbutamol and the injection marker oxprenolol. The injected sample plugs were separated by plugs of background electrolyte, through application of a short-term voltage (30 kV) over the capillary for different time periods, i.e., tPE1 and tPE2. The samples in each set were isolated from each other by partial electrophoresis for 2.35 min (tPE1), while the sample sets were separated for 10.50 min (tPE2). After the final injection, all the applied samples were subjected to electrophoresis for a time period corresponding to that in conventional single-injection CZE. The method was validated regarding linearity, accuracy, precision and robustness before it was applied to the determination of salbutamol in 15 tablets of Ventoline Depot with a labeled content of 8 mg salbutamol. The average salbutamol content was determined to 7.8 mg (±0.3 mg) from simultaneous analyses of the 15 different tablets.

  • 44.
    Lu, Lili
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
    Lundqvist, Andreas
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
    Zeng, Cheng-Ming
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
    Lagerquist, Christine
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
    Lundahl, Per
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry.
    D-Glucose, forskolin and cytochalasin B affinities for the glucose transporter Glut1. Study of pH and reconstitution effects by biomembrane affinity chromatography1997In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 776, no 1, p. 81-86Article in journal (Refereed)
    Abstract [en]

    The affinities of D-glucose and the transport inhibitors, forskolin and cytochalasin B (CB), for Glut1 were studied by frontal affinity chromatography at pH 5-10 on sterically immobilized proteoliposomes with reconstituted human red cell glucose transporter Glut1. The affinity of D-glucose for Glut1 became slightly weaker as the pH was increased. The inhibitor affinities decreased and became immeasurably weak above pH 9. At pH 7.4, the dissociation constants were 44 mM for glucose, 1.8 microM for forskolin and 72 nM for CB. The affinities of these solutes for Glut1 in red cell membrane vesicles and particularly for Glut1 in red cells were higher, as shown by chromatographic analyses.

  • 45.
    Lönnberg, Maria
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Carlsson, Jan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Lab-on-a-chip technology for determination of protein isoform profiles2006In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1127, no 1-2, p. 175-182Article in journal (Refereed)
    Abstract [en]

    A novel lab-on-a-chip technique for rapid (<15 min) and quantitative isoform-profile determination is presented. Ion-exchange chromatographic separation of protein-isoforms and a sensitive immunoassay detection are combined in a porous monolith chip. Thin lines of immobilized antibodies are used for specific capturing of target molecules, which can be detected by the reaction with antibodies bound to carbon black nano-strings. The bound carbon black is quantified by the use of an image scanner. As demonstrated with transferrin isoforms, differing only by 0.1 pH unit in their pI, this technology can distinguish minor differences in protein carbohydrate structure and enable specific determination of proteins in a complex environment, requiring only a few picogram of isoform for detection.

  • 46.
    Lönnberg, Maria
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Dehnes, Yvette
    Drevin, Malin
    Garle, Mats
    Lamon, Severine
    Leuenberger, Nicolas
    Quach, Trikien
    Carlsson, Jan
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Surface Biotechnology.
    Rapid affinity purification of erythropoietin from biological samples using disposable monoliths2010In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1217, no 45, p. 7031-7037Article in journal (Refereed)
    Abstract [en]

    Identification of post-translational modifications of proteins in biological samples often requires access to preanalytical purification and concentration methods In the purification step high or low molecular weight substances can be removed by size exclusion filters and high abundant proteins can be removed or low abundant proteins can be enriched by specific capturing tools In this paper is described the experience and results obtained with a recently emerged and easy-to-use affinity purification kit for enrichment of the low amounts of EPO found in urine and plasma specimens The kit can be used as a pre-step in the EPO doping control procedure as an alternative to the commonly used ultrafiltration for detecting aberrantly glycosylated isoforms The commercially available affinity purification kit contains small disposable anti-EPO monolith columns (6 mu L volume theta 7 mm length 0 15 mm) together with all required buffers A 24-channel vacuum manifold was used for simultaneous processing of samples The column concentrated EPO from 20 mL urine down to 55 mu L eluate with a concentration factor of 240 times while roughly 997% of non-relevant urine proteins were removed The recoveries of Neorecormon (epoetin beta) and the EPO analogues Aranesp and Mircera applied to buffer were high 76% 67% and 57% respectively The recovery of endogenous EPO from human urine was 65% High recoveries were also obtained when purifying human mouse and equine EPO from serum and human EPO from cerebrospinal fluid Evaluation with the accredited EPO doping control method based on isoelectric focusing (IEF) showed that the affinity purification procedure did not change the isoform distribution for rhEPO Aranesp Mircera or endogenous EPO The kit should be particularly useful for applications in which it is essential to avoid carry-over effects a problem commonly encountered with conventional particle-based affinity columns The encouraging results with EPO propose that similar affinity monoliths with the appropriate antibodies should constitute useful tools for general applications in sample preparation not only for doping control of EPO and other hormones such as growth hormone and insulin but also for the study of post-translational modifications of other low abundance proteins in biological and clinical research and for sample preparation prior to in vitro diagnostics.

  • 47.
    Mohabbati, Sheila
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Westerlund, Douglas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Improved properties of the non-covalent coating with N,N-didodecyl-N, N-dimethylammonium bromide for the separation of basic proteins by capillary electrophoresis with acidic buffers in 25mum capillaries2006In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1121, no 1, p. 32-39Article in journal (Refereed)
    Abstract [en]

    Capillaries (25 μm I.D.) treated with the double-alkyl-chain cationic surfactant N,N-didodecyl-N, N-dimethylammonium bromide (DDAB) in an improved coating procedure were used for separation of four basic proteins in volatile buffers (ammonium acetate and ammonium hydroxyacetate) as well as in a non-volatile buffer (sodium phosphate) at pH 4. The DDAB coating was stable enough to, without recoating, permit consecutive separations of the proteins up to 9 h with good precisions in peak areas (RSD = 1.1%) and migration times and with high apparent efficiencies (over 1 million theoretical plates/m) in the presence of a strong anodic electroosmosis. Adsorption of the proteins onto the capillary surface, which in previous studies was found to give a certain contribution to zone broadening, was eliminated with the new modified coating method. Complex formation between the proteins and phosphate buffer was studied and confirmed, and it is proposed that slow protein–buffer component interactions are the main contributions to zone broadening in protein separations by CE.

  • 48. Mu, H L
    et al.
    Wesen, C
    Novak, T
    Sundin, Peter
    Skramstad, J
    Odham, G
    Enrichment of chlorinated fatty acids in fish lipids prior to analysis by capillary gas chromatography with electrolytic conductivity detection and mass spectrometry1996In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 731, no 1-2, p. 225-236Article in journal (Refereed)
  • 49.
    Muszynska, Grazyna
    et al.
    Polish Academy of Science, Warsawa.
    Dobrowolska, Grazyna
    Polish Academy of Science, Warsawa.
    Medin, Anders
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
    Ekman, Pia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Porath, Jerker
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC, Biochemistry.
    Model studies on iron (III) ion affinity chromatography: II. Interaction of immobilized ferric ions with phosphorylated amino acids, peptides and proteins1992In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 604, no 1, p. 19-28Article in journal (Refereed)
    Abstract [en]

    The chromatographic behaviour of phosphoamino acids, phosphopeptides and phosphoproteins and their non-phosphorylated counterparts was studied on Fe(III)-Chelating Sepharose and Fe(III)-Chelating Superose. The phosphorylated compounds, in contrast to their non-phosphorylated or dephosphorylated counterparts, adsorb to immobilized iron(III) ions at pH 5.5 and can be desorbed by an increase in pH. Phosphoamino acids were eluted at pH 6.5-6.7, whereas monophosphopeptides and phosphoprotamine eluted in the pH range 6.9-7.5. Molecules possessing clusters(s) of carboxylic groups are weakly retained (gamma-carboxyglutamic acid, Ala-Ser-Glu5) or bound (polyglutamic acid, beta-casein) to the immobilized iron(III) ions at pH 5.5. Dephosphorylated beta-casein was desorbed at pH 7.0, whereas for elution of native (non-dephosphorylated) beta-casein, phosphate buffer of pH 7.7 was required. The homopolymer of polyglutamic acid was desorbed in the pH range 6.0-6.3, whereas copolymers of glutamic acid and tyrosine require pH 7.0-7.3 or even phosphate buffer at pH 7.7 for elution.

  • 50. Ovesen, Rikke G.
    et al.
    Göransson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Hansen, Steen H.
    Nielsen, John
    Hansen, Hans Christian B.
    A liquid chromatography-electrospray ionization-mass spectrometry method for quantification of cyclotides in plants avoiding sorption during sample preparation2011In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1218, no 44, p. 7964-7970Article in journal (Refereed)
    Abstract [en]

    Cyclotides are plant-produced, bioactive, cyclic mini-proteins with interesting pharmaceutical and agricultural applications. A reverse phase liquid chromatography electrospray ionization mass spectrometry (RP-LC-ESI-MS) method for analysis of cyclotides in plant materials with a minimum of sample pretreatment is presented. Three exemplary cyclotides (kalata B1, kalata B2 and cycloviolacin 02) were used as reference substances for the method development. Linearity (r(2) > 0.99) was achieved in the concentration range 0.05-10 mg/L and the limit of detection was 1.7-4.0 mu g/L. The present study is the first to demonstrate that cyclotides dissolved in water sorb to glass vials, but the addition of 15% of acetonitrile or 40 mg/L of bovine serum albumin is sufficient to keep the cyclotides in solution. Cyclotides were extracted from candied violets, violet tea, and the plants Oldenlandia affinis and Viola odorata using 70% methanol containing 0.1% formic acid (v/v). The plant content was determined to be 23.5-14,200 mu g/g (dry weight). The highest content of cyclotide was found in wild Danish V. odorata, and it is the highest content of cyclotide in a plant reported hitherto. Candied violets contained 0.00-8.66 mu g/g (dry weight), while no cyclotides were detected in commercial violet tea.

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