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  • 1.
    Aftab, Obaid
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    Engskog, Mikael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Elmsjö, Albert
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Hammerling, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    Gustafsson, Mats
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Cancer Pharmacology and Computational Medicine.
    NMR spectroscopy based metabolic profiling of drug induced changes in vitro can discriminate between pharmacological classes2014In: Journal of chemical information and modeling, ISSN 1549-9596, Vol. 54, no 11, p. 3251-3258Article in journal (Refereed)
    Abstract [en]

    Drug induced changes in mammalian cell line models have already been extensively profiled at the systemic mRNA level and subsequently used to suggest mechanisms of action for new substances as well as to support drug repurposing, i.e. identifying new potential indications for drugs already licensed for other pharmacotherapy settings. The seminal work in this field, which includes a large database and computational algorithms for pattern matching, is known as the “Connectivity Map” (CMap). The potential of similar exercises at the metabolite level is, however, still largely unexplored. Only recently the first high throughput metabolomic assay pilot study was published, involving screening of metabolic response to a set of 56 kinase inhibitors in a 96-well format. Here we report results from a separately developed metabolic profiling assay, which leverages 1H NMR spectroscopy to the quantification of metabolic changes in the HCT116 colorectal cancer cell line, in response to each of 26 compounds. These agents are distributed across 12 different pharmacological classes covering a broad spectrum of bioactivity. Differential metabolic profiles, inferred from multivariate spectral analysis of 18 spectral bins, allowed clustering of most tested drugs according to their respective pharmacological class. A more advanced supervised analysis, involving one multivariate scattering matrix per pharmacological class and using only 3 spectral bins (three metabolites), showed even more distinct pharmacology-related cluster formations. In conclusion, this kind of relatively fast and inexpensive profiling seems to provide a promising alternative to that afforded by mRNA expression analysis, which is relatively slow and costly. As also indicated by the present pilot study, the resulting metabolic profiles do not seem to provide as information rich signatures as those obtained using systemic mRNA profiling, but the methodology holds strong promise for significant refinement.

  • 2.
    Amini, Ahmad
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Lodén, Henrik
    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.
    Principles for different modes of multiple-injection CZE2008In: Electrophoresis, ISSN 0173-0835, E-ISSN 1522-2683, Vol. 29, no 19, p. 3952-3958Article in journal (Refereed)
    Abstract [en]

    This paper introduces four different modes of multiple-injection CZE (MICZE). The validity of these MICZE models was evaluated by the experimental data. Prior to the application of MICZE, the electrophoretic conditions are developed in the single-injection mode by adjusting different experimental parameters such as pH, type and concentration of buffer additives and temperature. Based on the migration time difference (tmig) between the analyte and the internal standard or injection marker, one or more MICZE modes can be employed. The injection marker is added to the sample to compensate for injection-volume fluctuations. The inter-plug distance is regulated by applying an electrical field over the capillary for a short period of time between each injection. After the final injection, the separation is completed by electrophoresis for a time period corresponding to that in the single-injection mode

  • 3.
    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.

  • 4.
    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.

  • 5.
    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.

  • 6.
    Carlsson, Y
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Hedeland, M
    Bondesson, U
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Pettersson, C
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Non-aqueous capillary electrophoretic separation of enantiomeric amineswith (-)-2,3:4,6-di-O-isopropylidene-2-keto-L-gulonic acid as chiralcounter ion.2001In: J Chromatogr A, Vol. 922, p. 303-Article in journal (Refereed)
  • 7.
    Carlsson, 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.
    Non-aquoeus capillary electrophoretic separation of enantiomeric amines with (-)-2,3:4,6-di-O-isopropylidene-2-keto-L-gulonic acid as chiral counter ion2001In: Journal of Chromatography A, ISSN 0021-9673, Vol. 922, no 1-2, p. 303-311Article in journal (Refereed)
    Abstract [en]

    (2)-2,3:4,6-Di-O-isopropylidene-2-keto-L-gulonic acid [(2)-DIKGA] has been introduced as a chiral counter ion innon-aqueous capillary electrophoresis. High enantioresolutions (R $3) were obtained for amines, e.g., pronethalol, labetalol Sand bambuterol. Methanol containing NaOH and (2)-DIKGA was used as the background electrolyte. The counter ionconcentration and the nature of the injection medium were found to affect the chiral separation. Covalent coating of thefused-silica capillary reduced the electro-osmotic flow resulting in improved enantioresolutions.

  • 8.
    Elmsjö, Albert
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Engskog, Mikael K. R.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Nestor, Marika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science.
    Arvidsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry. Med Prod Agcy, Uppsala, Sweden.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    The co-feature ratio, a novel method for the measurement of chromatographic and signal selectivity in LC-MS-based metabolomics.2017In: Analytica Chimica Acta, ISSN 0003-2670, E-ISSN 1873-4324, Vol. 956, p. 40-47Article in journal (Refereed)
    Abstract [en]

    Evaluation of analytical procedures, especially in regards to measuring chromatographic and signal selectivity, is highly challenging in untargeted metabolomics. The aim of this study was to suggest a new straightforward approach for a systematic examination of chromatographic and signal selectivity in LC-MS-based metabolomics. By calculating the ratio between each feature and its co-eluting features (the co-features), a measurement of the chromatographic selectivity (i.e. extent of co-elution) as well as the signal selectivity (e.g. amount of adduct formation) of each feature could be acquired, the co-feature ratio. This approach was used to examine possible differences in chromatographic and signal selectivity present in samples exposed to three different sample preparation procedures. The capability of the co-feature ratio was evaluated both in a classical targeted setting using isotope labelled standards as well as without standards in an untargeted setting. For the targeted analysis, several metabolites showed a skewed quantitative signal due to poor chromatographic selectivity and/or poor signal selectivity. Moreover, evaluation of the untargeted approach through multivariate analysis of the co-feature ratios demonstrated the possibility to screen for metabolites displaying poor chromatographic and/or signal selectivity characteristics. We conclude that the co-feature ratio can be a useful tool in the development and evaluation of analytical procedures in LC-MS-based metabolomics investigations. Increased selectivity through proper choice of analytical procedures may decrease the false positive and false negative discovery rate and thereby increase the validity of any metabolomic investigation.

  • 9.
    Elmsjö, Albert
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Rosqvist, Fredrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Clinical Nutrition and Metabolism.
    Engskog, Mikael K R
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Kullberg, Joel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Radiology.
    Iggman, David
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Clinical Nutrition and Metabolism.
    Johansson, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Radiology.
    Ahlström, Håkan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Radiology.
    Arvidsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Risérus, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Public Health and Caring Sciences, Clinical Nutrition and Metabolism.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    NMR-based metabolic profiling in healthy individuals overfed different types of fat: links to changes in liver fat accumulation and lean tissue mass.2015In: Nutrition & Diabetes, ISSN 2044-4052, E-ISSN 2044-4052, Vol. 5, no 19, p. e182-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Overeating different dietary fatty acids influence the amount of liver fat stored during weight gain, however, the mechanisms responsible are unclear. We aimed to identify non-lipid metabolites that may differentiate between saturated (SFA) and polyunsaturated fatty acid (PUFA) overfeeding using a non-targeted metabolomic approach. We also investigated the possible relationships between plasma metabolites and body fat accumulation.

    METHODS: In a randomized study (LIPOGAIN study), n=39 healthy individuals were overfed with muffins containing SFA or PUFA. Plasma samples were precipitated with cold acetonitrile and analyzed by nuclear magnetic resonance (NMR) spectroscopy. Pattern recognition techniques were used to overview the data, identify variables contributing to group classification and to correlate metabolites with fat accumulation.

    RESULTS: We previously reported that SFA causes a greater accumulation of liver fat, visceral fat and total body fat, whereas lean tissue levels increases less compared with PUFA, despite comparable weight gain. In this study, lactate and acetate were identified as important contributors to group classification between SFA and PUFA (P<0.05). Furthermore, the fat depots (total body fat, visceral adipose tissue and liver fat) and lean tissue correlated (P(corr)>0.5) all with two or more metabolites (for example, branched amino acids, alanine, acetate and lactate). The metabolite composition differed in a manner that may indicate higher insulin sensitivity after a diet with PUFA compared with SFA, but this needs to be confirmed in future studies.

    CONCLUSION: A non-lipid metabolic profiling approach only identified a few metabolites that differentiated between SFA and PUFA overfeeding. Whether these metabolite changes are involved in depot-specific fat storage and increased lean tissue mass during overeating needs further investigation.

  • 10.
    Engskog, Mikael K R
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Björklund, My
    Haglöf, Jakob
    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.
    Shoshan, Maria
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Metabolic profiling of epithelial ovarian cancer cell lines: evaluation of harvesting protocols for profiling using NMR spectroscopy2015In: Bioanalysis, ISSN 1757-6180, E-ISSN 1757-6199, Vol. 7, no 2, p. 157-166Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Metabolic profiling represents a novel technology for analyzing tumor cells. Epithelial ovarian carcinoma has a low survival rate due to the development of aggressive and chemotherapy-resistant cells. A tailored and reliable protocol is presented for profiling of chemoresistant cells using the cell line SKOV3 and a multiresistant subline SKOV3R.

    RESULTS: Harvesting protocols with cold methanol or MilliQ freeze/thaw cycles were compared. Increased reproducibility using MilliQ was evidenced. Importantly, both approaches resulted in similar profiles. Compared with parental SKOV3, the SKOV3R cells showed a significantly different profile.

    CONCLUSION: The MilliQ protocol is preferred owing to higher reproducibility and increased sample preparation options. The resulting metabolic profiles summarize metabolic alterations in chemoresistant cells consistent with a progressed and aggressive phenotype.

  • 11.
    Engskog, Mikael K. R.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Ersson, Lisa
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Haglöf, Jakob
    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. Medical Product Agency, Box 26, Dag Hammarskjölds väg 42, 751 03 Uppsala, Sweden.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Brittebo, Eva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    β-N-Methylamino-L-alanine (BMAA) perturbs alanine, aspartate and glutamate metabolism pathways in human neuroblastoma cells as determined by metabolic profiling2017In: Amino Acids, ISSN 0939-4451, E-ISSN 1438-2199, Vol. 49, no 5, p. 905-919Article in journal (Refereed)
    Abstract [en]

    β-Methylamino-L-alanine (BMAA) is a non-proteinogenic amino acid that induces long-term cognitive deficits, as well as an increased neurodegeneration and intracellular fibril formation in the hippocampus of adult rodents following short-time neonatal exposure and in vervet monkey brain following long-term exposure. It has also been proposed to be involved in the etiology of neurodegenerative disease in humans. The aim of this study was to identify metabolic effects not related to excitotoxicity or oxidative stress in human neuroblastoma SH-SY5Y cells. The effects of BMAA (50, 250, 1000 µM) for 24 h on cells differentiated with retinoic acid were studied. Samples were analyzed using LC-MS and NMR spectroscopy to detect altered intracellular polar metabolites. The analysis performed, followed by multivariate pattern recognition techniques, revealed significant perturbations in protein biosynthesis, amino acid metabolism pathways and citrate cycle. Of specific interest were the BMAA-induced alterations in alanine, aspartate and glutamate metabolism and as well as alterations in various neurotransmitters/neuromodulators such as GABA and taurine. The results indicate that BMAA can interfere with metabolic pathways involved in neurotransmission in human neuroblastoma cells.

  • 12.
    Engskog, Mikael K R
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Haglöf, Jakob
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Evaluation of harvesting protocols for metabolic profiling of epithelial ovarian cancer cells2014Conference paper (Other academic)
  • 13.
    Engskog, Mikael K. R.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Haglöf, Jakob
    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. Med Prod Agcy, Dag Hammarskjolds Vag 42,Box 26, SE-75103 Uppsala, Sweden..
    Pettersson, Curt
    LC-MS based global metabolite profiling: the necessity of high data quality2016In: Metabolomics, ISSN 1573-3882, E-ISSN 1573-3890, Vol. 12, no 7, article id 114Article, review/survey (Refereed)
    Abstract [en]

    LC-MS based global metabolite profiling currently lacks detailed guidelines to demonstrate that the obtained data is of high enough analytical quality. Insufficient data quality may result in the failure to generate a hypothesis, or in the worst case, a false or skewed hypothesis. After assessing the literature, it is apparent that an analytically focused summary and critical discussion related to this subject would be beneficial for both beginners and experts engaged in this field. A particular focus will be placed on data quality, which we here define as the degree to which a set of parameters fulfills predetermined criteria, similar to the established guidelines for targeted analysis. However, several of these parameters are difficult to assess since holistic approaches measure thousands of metabolites in parallel and seldom include predefined knowledge of which metabolites will differ between sample groups. In this review, the following parameters will be discussed in detail: reproducibility, selectivity, certainty of metabolite identification and metabolite coverage. The review systematically describes the generic workflow for LC-MS based global metabolite profiling and highlights how each separate part may affect data quality. The last part of the review describes how data quality can be evaluated as well as identifies areas where additional improvement is needed. In this review, we provide our own analytical opinions in regards to evaluation and, to some extent, improvement of data quality.

  • 14.
    Engskog, Mikael K R
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Karlsson, Oskar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Elmsjö, Albert
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Brittebo, Eva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Arvidsson, Torbjörn
    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.
    The cyanobacterial amino acid beta-N-methylamino-L-alanine perturbs the intermediary metabolism in neonatal rats2017In: Amino Acids, ISSN 0939-4451, E-ISSN 1438-2199, Vol. 49, no 5, p. 905-919, article id 10.1007/s00726-017-2391-8Article in journal (Refereed)
    Abstract [en]

    The neurotoxic amino acid β-N-methylamino-l-alanine (BMAA) is produced by most cyanobacteria. BMAA is considered as a potential health threat because of its putative role in neurodegenerative diseases. We have previously observed cognitive disturbances and morphological brain changes in adult rodents exposed to BMAA during the development. The aim of this study was to characterize changes of major intermediary metabolites in serum following neonatal exposure to BMAA using a non-targeted metabolomic approach. NMR spectroscopy was used to obtain serum metabolic profiles from neonatal rats exposed to BMAA (40, 150, 460mg/kg) or vehicle on postnatal days 9-10. Multivariate data analysis of binned NMR data indicated metabolic pattern differences between the different treatment groups. In particular five metabolites, d-glucose, lactate, 3-hydroxybutyrate, creatine and acetate, were changed in serum of BMAA-treated neonatal rats. These metabolites are associated with changes in energy metabolism and amino acid metabolism. Further statistical analysis disclosed that all the identified serum metabolites in the lowest dose group were significantly (p<0.05) decreased. The neonatal rat model used in this study is so far the only animal model that displays significant biochemical and behavioral effects after a low short-term dose of BMAA. The demonstrated perturbation of intermediary metabolism may contribute to BMAA-induced developmental changes that result in long-term effects on adult brain function.

  • 15.
    Fransson, Anette E
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Otolaryngology and Head and Neck Surgery.
    Kisiel, Marta
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Otolaryngology and Head and Neck Surgery.
    Pirttilä, Kristian
    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.
    Videhult Pierre, Pernilla
    Division of Audiology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden.
    Laurell, Göran
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Otolaryngology and Head and Neck Surgery.
    Hydrogen Inhalation Protects against Ototoxicity Induced by Intravenous Cisplatin in the Guinea Pig2017In: Frontiers in Cellular Neuroscience, ISSN 1662-5102, E-ISSN 1662-5102, Vol. 11, article id 280Article in journal (Refereed)
    Abstract [en]

    Introduction: Permanent hearing loss and tinnitus as side-effects from treatment with the anticancer drug cisplatin is a clinical problem. Ototoxicity may be reduced by co-administration of an otoprotective agent, but the results in humans have so far been modest.

    Aim: The present preclinical in vivo study aimed to explore the protective efficacy of hydrogen (H2) inhalation on ototoxicity induced by intravenous cisplatin.

    Materials and Methods: Albino guinea pigs were divided into four groups. The Cispt (n = 11) and Cispt+H2 (n = 11) groups were given intravenous cisplatin (8 mg/kg b.w., injection rate 0.2 ml/min). Immediately after, the Cispt+H2 group also received gaseous H2 (2% in air, 60 min). The H2 group (n = 5) received only H2 and the Control group (n = 7) received neither cisplatin nor H2. Ototoxicity was assessed by measuring frequency specific ABR thresholds before and 96 h after treatment, loss of inner (IHCs) and outer (OHCs) hair cells, and by performing densitometry-based immunohistochemistry analysis of cochlear synaptophysin, organic transporter 2 (OCT2), and copper transporter 1 (CTR1) at 12 and 7 mm from the round window. By utilizing metabolomics analysis of perilymph the change of metabolites in the perilymph was assessed.

    Results: Cisplatin induced electrophysiological threshold shifts, hair cell loss, and reduced synaptophysin immunoreactivity in the synapse area around the IHCs and OHCs. H2 inhalation mitigated all these effects. Cisplatin also reduced the OCT2 intensity in the inner and outer pillar cells and in the stria vascularis as well as the CTR1 intensity in the synapse area around the IHCs, the Deiters' cells, and the stria vascularis. H2 prevented the majority of these effects.

    Conclusion: H2 inhalation can reduce cisplatin-induced ototoxicity on functional, cellular, and subcellular levels. It is proposed that synaptopathy may serve as a marker for cisplatin ototoxicity. The effect of H2 on the antineoplastic activity of cisplatin needs to be further explored.

  • 16.
    Haglind, Alfred
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Hedeland, Mikael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry. National Veterinary Institute (SVA), Dept. of Chemistry, Environment and Feed Hygiene, Uppsala, Sweden.
    Arvidsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry. Medical Products Agency, Uppsala, Sweden.
    Pettersson, Curt E
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Major signal suppression from metal ion clusters in SFC/ESI-MS: Cause and Effects2018In: Journal of chromatography. B, ISSN 1570-0232, E-ISSN 1873-376X, Vol. 1084, p. 96-105Article in journal (Refereed)
    Abstract [en]

    The widening application area of SFC-MS with polar analytes and water-containing samples facilitates the use of quick and simple sample preparation techniques such as “dilute and shoot” and protein precipitation. This has also introduced new polar interfering components such as alkali metal ions naturally abundant in e.g. blood plasma and urine, which have shown to be retained using screening conditions in SFC/ESI-TOF-MS and causing areas of major ion suppression. Analytes co-eluting with these clusters will have a decreased signal intensity, which might have a major effect on both quantification and identification. When investigating the composition of the alkali metal clusters using accurate mass and isotopic pattern, it could be concluded that they were previously not described in the literature. Using NaCl and KCl standards and different chromatographic conditions, varying e.g. column and modifier, the clusters proved to be formed from the alkali metal ions in combination with the alcohol modifier and make-up solvent. Their compositions were [(XOCH3)n+X]+, [(XOH)n+X]+, [(X2CO3)n+X]+ and [(XOOCOCH3)n+X]+ for X= Na+ or K+ in ESI+. In ESI-, the clusters depended more on modifier, with [(XCl)n+Cl]- and [(XOCH3)n+OCH3]- mainly formed in pure methanol and [(XOOCH)n+OOCH]- when 20 mM NH4Fa was added.

    To prevent the formation of the clusters by avoiding methanol as modifier might be difficult, as this is a widely used modifier providing good solubility when analyzing polar compounds in SFC. A sample preparation with e.g. LLE would remove the alkali ions, however also introducing a time consuming and discriminating step into the method. Since the alkali metal ions were retained and affected by chromatographic adjustments as e.g. mobile phase modifications, a way to avoid them could therefore be chromatographic tuning, when analyzing samples containing them.

  • 17.
    Haglöf, Jakob
    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.
    Capillary electrophoretic studies of ion pairing in aqueous solution using N-derivatized dipeptides2008Conference paper (Other academic)
  • 18.
    Haglöf, Jakob
    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.
    Enantiomeric separation of amines using divalent N-derivatized dipeptides as chiral selectors in non-aqueous capillary electrophoresis2008Conference paper (Other academic)
  • 19.
    Haglöf, Jakob
    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.
    Separation of amino alcohols using divalent dipeptides as counter ions in aqueous CE2010In: Electrophoresis, ISSN 0173-0835, E-ISSN 1522-2683, Vol. 31, no 10, p. 1706-1712Article in journal (Refereed)
    Abstract [en]

    Divalent dipeptides have been introduced as counter ions in aqueous CZE. The dipeptides form ion pairs with amino alcohols in the BGE and facilitate the separation of amino alcohols. High concentrations of dipeptide caused reversed effective mobility for the analytes. The net charge of the dipeptide can be controlled using a buffer or a strong base, and regulates the interaction between the dipeptide and the amino alcohol. A stronger interaction and higher selectivity of amino alcohols was observed when the dipeptides were used as divalent counter ions, than in monovalent or uncharged form. Association constants for ion pairs between divalent dipeptides and amino alcohols can be used to enhance selectivity for amino alcohols in CZE. No chiral separation of amino alcohols was observed when using the dipeptides as ion-pairing chiral selectors in aqueous BGE, but addition of methanol to the BGE promoted enantioselectivity.

  • 20.
    Hedeland, Ylva
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Beronius, Per
    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.
    Effect of alkali metal hydroxides on the enantioseparation of amines using di-O-isopropylidene-keto-L-gulonic acid as the selector in NACE2006In: Electrophoresis, ISSN 0173-0835, E-ISSN 1522-2683, Vol. 27, no 22, p. 4469-4479Article in journal (Refereed)
    Abstract [en]

    The present work demonstrates the importance of the ionic composition in the BGE for enantioseparation. (-)-2,3:4,6-di-O-Isopropylidene-2-keto-L-gulonic acid ((-)-DIKGA) has been used as the chiral selector in methanolic and ethanolic BGEs. The influence of added alkali metal hydroxides on the EOF and the chiral separation of amines; (atenolol, isoprenaline, pindolol and propranolol) have been studied. The ion-pair formation constants in ethanol were determined by precision conductometry for the enantiomers of pindolol with (-)-DIKGA, for Li+, Na+ and Cs+ with (-)-DIKGA, and also for the corresponding alkali metal hydroxides. The effective mobilities and the enantiomeric mobility differences were affected by the type of alkali metal hydroxide (LiOH, NaOH, KOH, RbOH or CsOH) added to the BGE. The effective mobility and mobility difference were increased with decrease in solvated radius of the alkali metal cation. These differences could partly be correlated to the ion-pair formation constants of the alkali metal cations with the chiral selector, affecting the equilibrium concentration of the free selector. The electroosmosis was also affected by the alkali metal hydroxide added to the BGE. The cathodic electroosmosis decreased with decreasing solvated radius of the alkali metal cation added to the BGE. Interestingly, the cathodic EOF was even reversed, i.e. became anodic in the ethanolic BGEs containing KOH, RbOH or CsOH and the methanolic ones with RbOH and CsOH.

  • 21.
    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.

  • 22.
    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.

  • 23.
    Hedeland, Ylva
    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.
    Chiral Separations in Nonaqueous Media2010In: Chiral Separations by Capillary Electrophoresis / [ed] Ann van Eeckhaut, Yvette Michotte, Boca Raton: CRC Press, 2010, 1.uppl., p. 271-312Chapter in book (Other academic)
  • 24.
    Hedeland, Ylva
    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.
    HPLC In Chiral Pharmaceutical Analysis2011In: Handbook of HPLC / [ed] Danilo Corradini, Boca Raton: CRC Press, 2011, 2nd ed., p. 507-534Chapter in book (Other academic)
  • 25.
    Hellqvist, Alexander
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Electroosmotic markers2012Conference paper (Refereed)
  • 26.
    Hellqvist, Alexander
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Electroosmotic markers2012Conference paper (Refereed)
  • 27.
    Hellqvist, Alexander
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Evaluation of electroosmotic markers in aqueous and nonaqueous capillary electrophoresis2013In: Electrophoresis, ISSN 0173-0835, E-ISSN 1522-2683, Vol. 34, no 24, p. 3252-3259Article in journal (Refereed)
    Abstract [en]

    The most common method to determine the EOF in CE is to measure the migration time for a neutral marker. In this study, 12 compounds (three novel and some previously used) were investigated as EOF markers in aqueous and nonaqueous BGEs. In the aqueous buffer systems (ammonium acetate, sodium phosphate, and sodium borate) the evaluation included a wide pH range (2-12). Two BGEs contained chiral selectors (sulphated-β-CD, (-)-diketogulonic acid) and one that contained a micellar agent (SDS) were included in the study. The majority of the evaluated compounds were found to migrate with the EOF in the water-based BGEs and are thus useful as EOF markers. However, in the SDS-based BGE only four of the compounds (acetone, acrylamide, DMSO, and ethanol) were found to be applicable. In the nonaqueous BGEs 11 markers (acetone, acetophenone, acrylamide, anthracene, benzene, 4-(4-methoxybenzylamino)-7-nitro-2,1,3-benzoxadiazole, benzyl alcohol, 2,5-diphenyloxazole, ethanol, flavone, and mesityl oxide) seemed to be functional as EOF markers. Even though several of the evaluated compounds can be used as EOF markers in the investigated BGEs, the authors would recommend the use of acrylamide as a general marker for UV detection. Furthermore, the four fluorescent markers (of which three were novel) gave RSD values equal to the other markers and can be used for the determination of the EOF in CE or microchip CE with fluorescence detection.

  • 28.
    Häggblad Sahlberg, Sara
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Mortensen, Anja C.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Engskog, Mikael K. R.
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Glimelius, Bengt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Experimental and Clinical Oncology.
    Stenerlöw, Bo
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science.
    Nestor, Marika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medical Radiation Science. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Different functions of AKT1 and AKT2 in molecular pathways, cell migration and metabolism in colon cancer cells2017In: International Journal of Oncology, ISSN 1019-6439, Vol. 50, no 1, p. 5-14Article in journal (Refereed)
    Abstract [en]

    AKT is a central protein in many cellular pathways such as cell survival, proliferation, glucose uptake, metabolism, angiogenesis, as well as radiation and drug response. The three isoforms of AKT (AKT1, AKT2 and AKT3) are proposed to have different physiological functions, properties and expression patterns in a cell type-dependent manner. As of yet, not much is known about the influence of the different AKT isoforms in the genome and their effects in the metabolism of colorectal cancer cells. In the present study, DLD-1 isogenic AKT1, AKT2 and AKT'/2 knockout colon cancer cell lines were used as a model system in conjunction with the parental cell line in order to further elucidate the differences between the AKT isoforms and how they are involved in various cellular pathways. This was done using genome wide expression analyses, metabolic profiling and cell migration assays. In conclusion, downregulation of genes in the cell adhesion, extracellular matrix and Notch-pathways and upregulation of apoptosis and metastasis inhibitory genes in the p53-pathway, confirm that the knockout of both AKT1 and AKT2 will attenuate metastasis and tumor cell growth. This was verified with a reduction in migration rate in the AKT1 KO and AKT2 KO and most explicitly in the AKT1/2 KO. Furthermore, the knockout of AKT1, AKT2 or both, resulted in a reduction in lactate and alanine, suggesting that the metabolism of carbohydrates and glutathione was impaired. This was further verified in gene expression analyses, showing downregulation of genes involved in glucose metabolism. Additionally, both AKT1 KO and AKT2 KO demonstrated an impaired fatty acid metabolism. However, genes were upregulated in the Wnt and cell proliferation pathways, which could oppose this effect. AKT inhibition should therefore be combined with other effectors to attain the best effect.

  • 29.
    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.

  • 30.
    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.

  • 31.
    Niklison-Chirou, Maria Victoria
    et al.
    Queen Mary Univ London, Barts & London Sch Med & Dent, Blizard Inst, London E1 2AT, England..
    Erngren, Ida
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Engskog, Mikael K R
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Picard, Daniel
    Heinrich Heine Univ Dusseldorf, Dept Pediat Oncol Hematol & Clin Immunol, D-40225 Dusseldorf, Germany.;Heinrich Heine Univ Dusseldorf, Dept Neuropathol, Med Fac, D-40225 Dusseldorf, Germany.;German Canc Consortium DKTK, German Canc Res Ctr DKFZ, Dept Pediat Neurooncogen, D-69120 Heidelberg, Germany..
    Remke, Marc
    Heinrich Heine Univ Dusseldorf, Dept Pediat Oncol Hematol & Clin Immunol, D-40225 Dusseldorf, Germany.;Heinrich Heine Univ Dusseldorf, Dept Neuropathol, Med Fac, D-40225 Dusseldorf, Germany.;German Canc Consortium DKTK, German Canc Res Ctr DKFZ, Dept Pediat Neurooncogen, D-69120 Heidelberg, Germany..
    McPolin, Phelim Hugh Redmond
    Queen Mary Univ London, Barts & London Sch Med & Dent, Blizard Inst, London E1 2AT, England..
    Selby, Matthew
    Newcastle Univ, Wolfson Childhood Canc Res Ctr, Northern Inst Canc Res, Newcastle Upon Tyne NE1 7RU, Tyne & Wear, England..
    Williamson, Daniel
    Newcastle Univ, Wolfson Childhood Canc Res Ctr, Northern Inst Canc Res, Newcastle Upon Tyne NE1 7RU, Tyne & Wear, England..
    Clifford, Steven C.
    Newcastle Univ, Wolfson Childhood Canc Res Ctr, Northern Inst Canc Res, Newcastle Upon Tyne NE1 7RU, Tyne & Wear, England..
    Michod, David
    UCL, Inst Child Hlth, London WC1N 1EH, England..
    Hadjiandreou, Michalis
    Queen Mary Univ London, Barts & London Sch Med & Dent, Blizard Inst, London E1 2AT, England..
    Arvidsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry. Med Prod Agcy, SE-75103 Uppsala, Sweden..
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Melino, Gerry
    Univ Leicester, MRC, Toxicol Unit, Leicester LE1 9HN, Leics, England..
    Marino, Silvia
    Queen Mary Univ London, Barts & London Sch Med & Dent, Blizard Inst, London E1 2AT, England..
    TAp73 is a marker of glutamine addiction in medulloblastoma2017In: Genes & Development, ISSN 0890-9369, E-ISSN 1549-5477, Vol. 31, no 17, p. 1738-1753Article in journal (Refereed)
    Abstract [en]

    Medulloblastoma is the most common solid primary brain tumor in children. Remarkable advancements in the understanding of the genetic and epigenetic basis of these tumors have informed their recent molecular classification. However, the genotype/phenotype correlation of the subgroups remains largely uncharacterized. In particular, the metabolic phenotype is of great interest because of its druggability, which could lead to the development of novel and more tailored therapies for a subset of medulloblastoma. p73 plays a critical role in a range of cellular metabolic processes. We show overexpression of p73 in a proportion of non-WNT medulloblastoma. In these tumors, p73 sustains cell growth and proliferation via regulation of glutamine metabolism. We validated our results in a xenograft model in which we observed an increase in survival time in mice on a glutamine restriction diet. Notably, glutamine starvation has a synergistic effect with cisplatin, a component of the current medulloblastoma chemotherapy. These findings raise the possibility that glutamine depletion can be used as an adjuvant treatment for p73-expressing medulloblastoma.

  • 32.
    Persson, Anita M.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Baumann, Kajsa
    Inst för kemi, Göteborgs universitet.
    Sundelöf, Lars-Olof
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Organic Pharmaceutical Chemistry.
    Lindberg, Walter
    AstraZeneca, Mölndal.
    Sokolowski, Anders
    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.
    Design and Characterization of a New Miniaturized Rotating Disk Equipment for In Vitro Dissolution Rate Studies2008In: Journal of Pharmaceutical Sciences, ISSN 0022-3549, E-ISSN 1520-6017, Vol. 97, no 8, p. 3344-3355Article in journal (Refereed)
    Abstract [en]

    A miniaturized apparatus for the determination of the apparent in vitro dissolution rate has been designed, constructed and characterized. The miniaturized apparatus was based on a low volume dissolution cell and a disk in a rotating magnetic bar. The disk tablet is pressed directly into the bar with a press designed and constructed for this purpose. It requires approximately 5 mg of substance. The disk was positioned eccentrically on the bar with an external flow of medium to increase the rate of solvent flow over the disk surface. Six different drug substances were used. The dissolution media were sodium phosphate buffer, pH 7.0, and ammonium acetate buffer, pH 6.8. All quantifications were made by integrating the dissolution cell with high-performance liquid chromatography (HPLC) using diode-array detection (DAD). The obtained results were compared with data from a conventional rotating disk equipment, where the disk was centrically mounted. The dissolution rates at 100 rpm seemed to be on an average of 2-3 times higher for the miniaturized apparatus (RSD 0.2-56%). The preliminary studies of this prototype indicate that the miniaturized rotating disk is a promising design for the qualitative estimation of dissolution rates of substances, for example during screening in early drug discovery.

  • 33.
    Persson, Anita M.
    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.
    Sokolowski, Anders
    AS Consulting, Uppsala.
    Correlation of in vitro dissolution rate and apparent solubility in buffered media using a miniaturized rotating disk equipment: Part II. Comparing different buffer media2009In: Drug Discoveries & Therapeutics, ISSN 1881-7831, Vol. 3, no 3, p. 114-122Article in journal (Refereed)
    Abstract [en]

    A correlation of the logarithmic values of the in vitro dissolution rate, G, and apparent solubility, S, was made for seven different drug substances from all of the classes in the Biopharmaceutics Classification System (BCS), in four different phosphate buffers. The effect of inorganic salts added as sodium chloride, sodium nitrate, sodium phosphate and sodium sulfate in the buffer media was investigated for the correlation. Triethanolammonium acetate buffer was also included in the study of the correlation of logG vs. logS. The pH was 7.0 ± 0.1 in all of the buffers to mimic a pH condition in intestinal fluids. The dissolution rate was determined with a newly constructed miniaturized rotating disk equipment, which enables fast determinations and consumes only minute quantities of substance (about 5 mg). The solubility was determined by conventional shake-flask methodology, using 1.5 mL solution volumes. All quantifications were performed with reversed phase high-performance liquid chromatography (RP-HPLC) and diode array detection (DAD). The different inorganic anions seemed to affect the solubility more than the dissolution rate. The phosphate and nitrate ions decreased the solubility for amines compared to the chloride ion. The best correlations of logG and logS were however obtained with a triethanolammonium acetate buffer. The good correlation (R2 = 0.991) may be sufficient in initial screening of drug solubility, based on dissolution rates in aqueous buffer media.

  • 34.
    Persson, Anita M.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Sokolowski, Anders
    AS Consulting, Uppsala.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Correlation of in vitro dissolution rate and apparent solubility in buffered media using a miniaturized rotating disk equipment: Part I. Comparison with a traditional USP rotating disk apparatus2009In: Drug Discoveries & Therapeutics, ISSN 1881-7831, Vol. 3, no 3, p. 104-113Article in journal (Refereed)
    Abstract [en]

    A correlation of the logarithmic values of the in vitro dissolution rate, G, and the apparent solubility, S, was evaluated in phosphate and ammonium acetate buffer at an initial pH of 7. The dissolution rates were determined with a newly designed and build miniaturized rotating disk equipment, as well as with a traditional rotating disk apparatus. The two apparatuses gave the same correlation pattern of logG and logS. Thirteen diverse drug substances from all of the classes in the Biopharmaceutics Classification System (BCS) were used for the correlation in the phosphate buffer system, with the results from the miniaturized apparatus only. A coefficient of determination, R2, of 0.982 was found if bases formulated as hydrochloride salts were excluded in the correlation. The miniaturized equipment is used for rapid screening of the dissolution rate, approximately 10 min for one run, and consumes small amounts of substance (about 5 mg) and dissolution media. All quantifications were performed by using reversed phase high-performance liquid chromatography (RPHPLC) with a diode array detector (DAD), integrated with the miniaturized rotating disk equipment.

  • 35.
    Persson, Anita Maria
    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.
    Rosén, Josefin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Multivariate data analysis of factors affecting the in vitro dissolution rate and the apparent solubility for a model basic drug substance in aqueous media2010In: Pharmaceutical research, ISSN 0724-8741, E-ISSN 1573-904X, Vol. 27, no 7, p. 1309-1317Article in journal (Refereed)
    Abstract [en]

    Purpose. To evaluate the usefulness of a miniaturized rotating disk equipment for the determination of factors influencing the in vitro dissolution rate, G, of a model basic drug substance (terfenadine) in different aqueous media, using experimental design and multivariate data analysis. The apparent solubility, S, was included in the chemometric study.

    Methods. The dissolution rate was determined with a miniaturized rotating disk apparatus and the solubility by shake-flask methodology. Media were based on acetate, phosphate or maleate buffers. The later used in fasted state simulated intestinal fluid (FaSSIF-V2). The chemometric analyses included fractional factorial design, principal component analysis (PCA) and orthogonal partial least squares (OPLS). Quantifications were made with a RP-HPLC-DAD system.

    Results. The most influential factor for both G and S of terfenadine in the different media was pH. Apart from the ionic strength and sodium chloride concentration in the acetate medium, the effects of the other variables were insignificant implying no wetting effect of the surfactants.

    Conclusions. The miniaturized rotating disk equipment was suitable to use, in conjunction with the chemometric analyses, in the evaluation of the factors affecting the in vitro dissolution rate. The apparent solubility was found to be influenced by the same factors as G.

  • 36. Stålberg, Olle
    et al.
    Kruusmägi, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Svensson, Mats A.
    Norinder, U.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
    Structure-interaction relationships between the bile acid GCA and pharmaceuticals using multivariate data analysis and capillary electrophoresis2007In: Journal of Pharmaceutical Sciences, ISSN 0022-3549, E-ISSN 1520-6017, Vol. 96, no 8, p. 2057-2073Article in journal (Refereed)
    Abstract [en]

    Capillary electrophoresis (CE) has been used in an interaction study of 66 pharmaceutical compounds with the bile acid glycocholate (GCA). The developed method proved to have a high precision in its ability to determine the mobility of drugs in buffer and buffer bile acids solutions. The relationship between solute structure and interaction with GCA was studied using two-dimensional descriptors with the in-house software SELMA and a three-dimensional model (quantum mechanical descriptors) in combination with the experimental CE-interaction data. The multivariate analysis method used was projection to latent structures by means of partial least squares (PLS). Two selections of training and test set were used for evaluation of a two-class model on interaction data. In the first selection all observations were used for training set, for example, creating a model, and re-predicting the observations on the model. A successful prediction on 85% of the drugs was observed using this model. The second selection used the 21 first tested compounds in the training set, where 78% of the compounds were correctly predicted using the two-dimensional model (SELMA) on the remaining 45 compounds and, respectively, 82% using the three-dimensional (quantum mechanical) model. Analysis of the impact of the descriptors showed that descriptors relating to hydrophobicity have a large positive effect on the interaction. Descriptors relating to polar properties have a pronounced negative effect on the interaction of drugs with bile acids.

  • 37.
    Svan, Alfred
    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.
    Arvidsson, Torbjorn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Jasper, Justin T.
    Sedlak, David L.
    Pettersson, Curt E.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Rapid chiral separation of atenolol, metoprolol, propranolol and the zwitterionic metoprolol acid using supercritical fluid chromatography-tandem mass spectrometry - Application to wetland microcosms2015In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1409, p. 251-258Article in journal (Refereed)
    Abstract [en]

    A method for enantiomeric separation of the three beta-blocking agents atenolol, metoprolol, propranolol and the zwitterionic metoprolol acid, a major metabolite of both metoprolol and in environmental matrices also atenolol, has been developed. By use of supercritical fluid chromatography and the polysaccharide-based Chiralpak (R) IB-3, all four compounds were simultaneously enantiomerically separated (R-s >1.5) within 8 min. Detection was performed using tandem mass spectrometry, and to avoid isobaric interference between the co-eluting metoprolol and metoprolol acid, the achiral column Acquity (R) UPC2 BEH 2-EP was attached ahead of to the chiral column. Carbon dioxide with 18% methanol containing 0.5% (v/v) of the additives trifluoroacetic acid and ammonia in a 2:1 molar ratio were used as mobile phase. A post column make-up flow (0.3 mL/min) of methanol containing 0.1% (v/v) formic acid was used to enhance the positive electrospray ionization. Detection was carried out using a triple quadrupole mass spectrometer operating in the selected reaction monitoring mode, using one transition per analyte and internal standard. The method was successfully applied for monitoring the enantiomeric fraction change over time in a laboratory scale wetland degradation study. It showed good precision, recovery, sensitivity and low effect of the sample matrix.

  • 38.
    Svan, Alfred
    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. Statens veterinärmedicinska anstalt (SVA) Enhet för kemi, miljö och fodersäkerhet.
    Arvidsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry. Läkemedelsverket.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Chiral separation of three β-blocking pharmaceuticals and a majormetabolite using SFC-MS2013Conference paper (Other academic)
  • 39.
    Svan, Alfred
    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.
    Arvidsson, Torbjörn
    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.
    Matrix effects: Do they differ between SFC/ESI-MS and LC/ESI-MS2016Conference paper (Other academic)
    Abstract [en]

    Introduction (120 ord)

    Supercritical fluid chromatography (SFC) is increasingly used in many fields following the new generation of instruments available on the market, often combined with electrospray ionization mass spectrometry (ESI/MS). New methods are developed and validated, often for samples of complex matrices. Thus, measurements of the matrix effects are sometimes included. Comparisons between SFC/ESI/MS and LC/ESI/MS have previously been performed, often focusing on e.g. sensitivity and separation. However, a systematic and qualitative investigation of the matrix effects achieved using same ion source and mass spectrometer, but different types of chromatography, i.e. SFC or LC, is so far lacking.

     

    Methods (120 ord)

    Effluent wastewater was collected and 100.0 ml was extracted using a generic SPE method (Oasis MCX), evaporated and redissolved in 1.0 ml of water:ACN (1:1). A 10 min gradient was developed for each technique, separating eight compounds chosen for being low molecular weight drugs with varying hydrophobicity and proteolytic properties. The mixture of compounds was then added to the ESI make-up solvent (MeOH) for the SFC/ESI/MS method. The extracted wastewater was injected without added compounds, and the SRM channel for each compound was monitored using tandem quadrupole MS (Quattro Micro, Waters®). The same interface and settings were used for LC, but an infusion of compounds was performed through a post-column syringe pump since no make-up flow was used.

     

    Preliminary data – Limit 300 words

    The retention times for the eight model compounds were evenly spread over the chromatographic time scale for both chromatographic methods. When applying the continuous compound infusion through the make-up solution (SFC/ESI/MS) or through a syringe pump (LC/ESI/MS), a stable signal was observed for all compounds. Injection of the extracted wastewater affected the signal in comparison with the blank. The difference in the signal profiles were however larger between LC and SFC than between blank and extracted wastewater. The two separation techniques also gave different levels of noise and variations in the SRM-channels for the different compounds, and occurred at differently time points during the gradients for both the techniques. In summary, the matrix effects seem to affect the detection differently depending on what chromatographic technique that is used. With increased knowledge about this, it could help future method development to minimize the matrix effects.    

     

    Novel aspect – Limit 20 words

    The first systematic comparison between SFC and LC in terms of matrix effects for ESI-MS/MS using qualitative methods (infusion).

  • 40.
    Svan, Alfred
    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. Natl Vet Inst SVA, Dept Chem Environm & Feed Hyg, SE-75189 Uppsala, Sweden..
    Arvidsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry. Med Prod Agcy, Box 26, SE-75103 Uppsala, Sweden..
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    The differences in matrix effect between supercritical fluid chromatography and reversed phase liquid chromatography coupled to ESI/MS2018In: Analytica Chimica Acta, ISSN 0003-2670, E-ISSN 1873-4324, Vol. 1000, p. 163-171Article in journal (Refereed)
    Abstract [en]

    For many sample matrices, matrix effects are a troublesome phenomenon using the electrospray ionization source. The increasing use of supercritical fluid chromatography with CO2 in combination with the electrospray ionization source for MS detection is therefore raising questions: is the matrix effect behaving differently using SFC in comparison with reversed phase LC? This was investigated using urine, plasma, influent-and effluent-wastewater as sample matrices. The matrix effect was evaluated using the post-extraction addition method and through post-column infusions. Matrix effect profiles generated from the post-column infusions in combination with time of flight-MS detection provided the most valuable information for the study. The combination of both qualitative and semi-quantitative information with the ability to use HRMS-data for identifying interfering compounds from the same experiment was very useful, and has to the authors' knowledge not been used this way before. The results showed that both LC and SFC are affected by matrix effects, however differently depending on sample matrix. Generally, both suppressions and enhancements were seen, with a higher amount of enhancements for LC, where 65% of all compounds and all sample matrices were enhanced, compared to only 7% for SFC. Several interferences were tentatively identified, with phospholipids, creatinine, and metal ion clusters as examples of important interferences, with different impact depending on chromatographic technique. SFC needs a different strategy for limiting matrix interferences, owing to its almost reverse retention order compared to RPLC.

  • 41.
    Svan, Alfred
    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.
    Arvidsson, Torbjörn
    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.
    The differences in matrix effects between supercritical fluid chromatography and reversed phase liquid chromatography coupled to ESI/MS analyzing blood plasma2017Conference paper (Other academic)
    Abstract [en]

    Introduction

    The increasing popularity of supercritical fluid chromatography (SFC) in combination with electrospray ionization mass spectrometry (ESI/MS) within several fields calls for a deeper knowledge regarding this combination of techniques. The ESI source is known for its sensitivity regarding matrix effects, often a factor controlled during method development and validation using LC. The different chemistry and chromatographic selectivity of LC and SFC give potentially different impact on the ionization process in ESI; however, this an area still not well studied.   

    Aim: To investigate how the matrix effects in ESI/MS differ for human plasma samples between SFC and reversed-phase LC, using generic screening conditions for both techniques, and a set of typical low molecular weight drug substances.

    Methods

    Pooled human plasma (500 µl) was precipitated using ice-cold acetonitrile (1000 µl). After mixing and centrifuging, 1200 µl of the supernatant were removed and evaporated at 40 ̊C. When dry, the samples were dissolved in 500µl water+0.1% FA (for LC) or acetonitrile:water 75:25 (for SFC). The samples were analyzed using SFC (Acquity UPC2, Waters®) and LC (Acquity UPLC, Waters®) and general screening conditions, using 10 min gradients. The same MS-system, a Q-ToF (Synapt G2-S, Waters®) acquiring in full scan mode, was used for detection with both separation techniques. The matrix effect was mainly evaluated using the Matrix Effect Profile, achieved from post-column compound infusions and injections of pretreated sample matrix and neat standards. From these data the average ME% was calculated for each data-point in the chromatogram, and through the full-scan mode using ToF, the compounds co-eluting with areas of suppression could be tentatively identified, suspected of creating the suppression. 

     

    Results and discussion The Matrix Effect Profile-evaluation of the experiments, combining qualitative and quantitative information with the added ability to use HRMS-data to identify interfering compounds from the same experiments were most useful for our aim. Phospholipids, creatinine, polyethylene glycol and cluster formations are examples of important interferences co-eluting with areas of ion suppression, but with different impact depending on chromatographic technique. The results also showed several areas of enhancement using LC, an effect not seen using SFC. 

  • 42.
    Sänger - van de Griend, Cari
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Capillary Electrophoresis: an Attractive Technique for Chiral Separations2013In: Chromatography Today, ISSN 1752-8070, Vol. 6, no 2, p. 32-37Article, review/survey (Refereed)
  • 43. Videhult Pierre, Pernilla
    et al.
    Haglöf, Jakob
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Linder, Birgitta
    Engskog, Mikael K R
    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.
    Pettersson, Curt
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Analytical Pharmaceutical Chemistry.
    Laurell, Göran
    Ototoxicity of Cisplatin Correlates with Changes in Blood Antioxidant Status2014Conference paper (Other academic)
1 - 43 of 43
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