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  • 51.
    Björling, Thomas
    Stockholm University, Faculty of Science, Department of Physical, Inorganic and Structural Chemistry.
    Synthesis and characterisation of Zintl hydrides2008Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The synthesis, structural characterisation and the properties of the Zintl hydrides AeE2H2 and AeAlSiH (Ae = Ba, Ca, Sr; E = Al, Ga, In, Si, Zn) are reported. The first hydride in this class of compounds is SrAl2H2 which was discovered under an experiment by Gingl, who hydrogenated SrAl2 at various temperatures. (Gingl et al, Journal of Alloys and Compounds 306 (2000) 127-132). The intention was to form alanates, e.g. AlH4-, by terminating the three dimensional four connected aluminium network in SrAl2. The new hydride, SrAl2H2, has a partially conserved aluminium network. The three dimensional anionic network in SrAl2 is reduced to two dimensions in the hydride, with aluminium bonded to both aluminium and hydrogen. This type of bonding configuration has not been observed before.

    The hydrogenation of SrAl2 is straight forward, 190 oC and 50 bar, compared to the difficult synthesis of alanates and alane, AlH3. The latter synthesises uses aluminium in its zero oxidation state in contrast to the synthesis of SrAl2H2 from SrAl2. (In the SrAl2-precursor aluminium is reduced by the electropositive metal to -I.) Thus, the discovery shows a different route to alanates by using precursors with aluminium in a reduced state. If SrAl2H2 is further hydrogenated at 250 oC the two dimensional network breaks and Sr2AlH7 forms.

    We wanted to investigate if SrAl2H2 was a singularity or if other similar compounds exist. We wanted to study how hydrogenation of precursors similar to the aluminide result in 1) new routes to compounds with high hydrogen content, as alanates, 2) to investigate how the E-H bond is affected as function of the network composition among different ternary hydrides, in particular BaAlxSi2-xHx, and choice of active metal.

    BaGa2H2 and SrGa2H2, two hydrides isostructural with SrAl2H2, were synthesized from its precursors BaGa2 and SrGa2. In addition three ternary hydrides BaAlSiH, CaAlSiH and SrAlSiH were manufactured from their related AeAlSi precursors.

    All powders were characterized by neutron and x-ray diffraction methods.

    An increased stability towards water/moisture compared to ordinary saline hydrides was noticed, especially for the ternary hydrides. Heat stability was measured with DSC (differential scanning calorimetry). The hydrides BaGa2H2 and SrGa2H2 decompose around 300 oC at 1 atm. This is similar to isostructural SrAl2H2. The ternary hydrides BaAlSiH and SrAlSiH decompose at 600 oC, at 1 atm, which is the highest noticed temperature for compounds with Al-H bonds. Inelastic neutron scattering experiments showed that these hydrides Al-H and Sr-H bonds are really weak, even weaker then the Al-H interactions in alanates and alanes. These hydrides are probably stabilized be their lattices. The electric properties among the ternary hydrides were measured with IR-spectroscopy (diffuse reflectance). The ternary hydrides, AeAlSiH, are indirect semi conductors. BaGa2H2 and SrGa2H2 are conductors. The ternary hydrides, AeAlxSi2-xHx, may have adjustable band gaps, which we were not able to determine.

    This work is leading into a new research area within the field of metal hydrides.

  • 52.
    Blom, Elisabeth
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Karimi, Farhad
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Eriksson, Olof
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology.
    Hall, Håkan
    Långström, Bengt
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Synthesis and in vitro evaluation of 18F-β-carboline alkaloids as PET ligands2008In: Journal of labelled compounds & radiopharmaceuticals, ISSN 0362-4803, E-ISSN 1099-1344, Vol. 51, no 6, p. 277-282Article in journal (Refereed)
    Abstract [en]

    A one-step 18F-labelling strategy was used to prepare four 18F-labelled analogues of 7-methoxy-1-methyl-9H-β-carboline (harmine): 7-(2-[18F]fluoroethoxy)-1-methyl-9H-β-carboline (5), 7-(3-[18F]fluoro-propoxy)-1-methyl-9H-β-carboline (6), 7-[2-(2-[18F]fluoroethoxy)ethoxy]-1-methyl-9H-β-carboline (7), and 7-{2-[2-(2-[18F]fluoroethoxy)ethoxy]-ethoxy}-1-methyl-9H-β-carboline (8). These were synthesized as potential PET ligands for monoamine oxidase A. A solution of pure labelled compound in buffer was obtained in < 70 min from end of radionuclide production, with a decay-corrected yield of up to 23%. The average specific binding to MAO-A in rat brain, determined by autoradiography experiments, was highest for compounds 7 and 8 (89 ± 2 and 96 ± 1% respectively), which was obtained at < 1 nM radioligand concentration.

  • 53.
    Blom, Elisabeth
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Karimi, Farhad
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Långström, Bengt
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    [F-18]/F-19 exchange in fluorine containing compounds for potential use in F-18-labelling strategies2009In: Journal of labelled compounds & radiopharmaceuticals, ISSN 0362-4803, E-ISSN 1099-1344, Vol. 52, no 12, p. 504-511Article in journal (Refereed)
    Abstract [en]

    Exchange of [F-18]fluoride with F-19 in various organofluorine compounds in concentrations ranging from 0.06 to 56 mM was explored. We aimed to explore whether exchange reactions can be a potential useful labelling strategy, when there are no requirement of high specific radioactivity. Parameters such as solvents, temperature, conventional vs microwave heating, and the degree of fluorine load in some aromatic and alkyl compounds were investigated with regard to radiochemical yield and specific radioactivity. A series of fluorobenzophenones (1-6), 1-(4-fluorophenyl)ethanone (7), various activated and deactivated fluoro benzenes (8-16), N-(pentafluorophenyl)benzamide (17), (pentafluorophenyl)formamide (18), (tridecafluorohexyl) benzene (19) and tetradecafluorohexane (20) were subjected to [F-18]/F-19 exchange. To test this strategy to label biologically active molecules containing fluorine atoms in an aryl group, two analogues of WAY-100635 (21-22), Lapatinib (23), 2,5,6,7,8-pentafluoro-3-methyinaphthoquinone (24) and 1-(2,4-difluorophenyl)-3-(4-fluorophenyl)propan-l-one (25) were investigated. The multi-fluorinated molecules containing an electron-withdrawing group were successfully labelled at room temperature, whereas the monofluorinated, as well as those containing an electron-donating group, required heating for the exchange reaction to take place.

  • 54.
    Blomqvist, Helen
    Stockholm University, Faculty of Science, Department of Physical, Inorganic and Structural Chemistry.
    Magnesium ions stabilizing solid-state transition metal hydrides2003Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis is focused on hydrides containing first-row transition metal–hydrido complexes counterbalanced by magnesium. In particular, fundamental properties of the promising hydrogen storage material Mg2NiH4 have been studied. Results presented in the thesis provide detailed knowledge of the electronic structure and bonding mechanisms and are of significance for improving the hydrogen desorption process of complex transition metal hydrides in general.Two competing stabilization mechanisms in Mg2NiH4 have been identified; presence of microtwinning in the structure and the extra Mg added to the melt-cast Mg2Ni starting alloys, which acts as stabilizing dopant in the Mg2NiH4 system. When eliminating both stabilization mechanisms, the hydrogen release pressure was doubled at 453 K. Mg2NiH4 behaves like a heavily doped semiconductor at lower temperatures, but the conductivity is counteracted by the introduction of microtwinning in the structure at elevated temperatures, which makes the hydride non-conducting at » 400 K. The conductivity can be regained by reducing the amount of microtwinning with an applied mechanical pressure. Size, coordination and type of cation framework have decisive roles in determining the structure type of complex metal hydrides, as shown by ab initio total-energy calculations. Compared to the rich variety of Pd-based complex hydrides, the few Ni hydrido complexes hitherto found only contain Mg2+, solely or in combination with Ca2+, Sr2+, Yb2+, Eu2+ or La3+. Apparently the rather weak Ni–H bond needs a small and polarizing cation, e.g. Mg2+, to be stabilized in the solid state. The rather strong Mg–H interaction makes Mg2NiH4 a hybrid of ionic and complex transition metal hydride. This stabilization mechanism where electron density is redistributed by the polarizable hydrido ligand explains why Mg2NiH4 is very sensitive to disturbances of the crystal ordering, e.g. doping and ball milling, which profoundly affect stability, conductivity, color, structure and phasetransition conditions. Mg2+ also has the ability to stabilize Mn hydrido complexes, as observed in the novel Mg3MnH~6 compound, synthesized at GPa pressure in the solid state.

  • 55.
    Blute, Irena
    et al.
    KTH, School of Chemical Science and Engineering (CHE), Chemistry.
    Pugh, Robert J.
    KTH, School of Chemical Science and Engineering (CHE), Chemistry.
    van de Pas, John
    Callaghan, Ian
    Industrial manufactured silica nanoparticle sols. 2: Surface tension, particle concentration, foam generation and stability2009In: Colloids and Surfaces A: Physicochemical and Engineering Aspects, ISSN 0927-7757, E-ISSN 1873-4359, Vol. 337, no 1-3, p. 127-135Article in journal (Refereed)
    Abstract [en]

    Several earlier papers have revealed that several key parameters, such as hydrophobicity (contact angle), size, shape and degree of agglomeration, have an important influence on the behavior of particles at the air/water interface. However, the origin of foaming with particles is still not clear. In this article, we have tentatively related surface tension measurements and particle concentrations to the generation and stability of foam produced from industrial manufactured silica nanoparticle sols. Surprisingly, only slight reductions in surface tension were observed and the differences between the hydrophophilic and partially modified hydrophobic sols were small. However, in the case of the partially modified hydrophobic sol, the surface tension/concentration gradient was found to be pH and concentration responsive. Also, the greatest reduction in Surface tension was found to occur at low pH (in the region of the pH(pzc)) and could be related to the highest foamability (foam generation) as determined in our earlier publication [1. Blute, R.J. Pugh, J. van de Pas, I. Callaghan, Silica nanoparticle sols. 1. Surface chemical characterization and evaluation of the foam generation (foamability), J. Colloid Interface Sci. 313 (2007) 645-655]. Also, after centrifugation of the moderately hydrophobic modified concentrated sols, foaming tests carried out on the supernatant indicated that the particle concentration had a dominant influence on foamability and foam stability. Since only transient foams, with relatively short lifetimes, could be produced with these modified silica nanoparticles then (a) further surface modification or the reduction of pH to increase the Surface activity or (b) the addition of a cosurfactant Would be needed to increase the foamability and achieve foams with extended lifetimes.

  • 56.
    Bodin, Rebecka
    Mälardalen University, School of Sustainable Development of Society and Technology.
    T-RFLP analyses of biocides influence on white water micro-organisms – planktonic and in biofilmIndependent thesis Advanced level (degree of Master (One Year)), 20 credits / 30 HE creditsStudent thesis
    Abstract [en]

    When paper is manufactured, deposits often form in the machines. These deposits are slimelike and can interfere with the papermaking process. The slimelike deposits are aggregates of micro-organisms, also known as biofilm. One single type of micro-organism can form a biofilm, but most biofilms consists of a mixture of several different kinds of micro-organisms and can form on about any conceivable surface. To control the aggregates of micro-organisms a slimecide is added, a so-called biocide. To examine what kind of bacteria that is included in the biofilm and also which bacteria that is killed or not killed by the biocide, Terminal Restriction Fragment Length Polymorphism analysis (T-RFLP) can be used.

     

    In this report we examine biocides impact on biofilm produced in the laboratory.The biocides were first tested for possible interference with the PCR-step of the T-RFLP analysis. None of the tested ten biocides inhibited the PCR process the biofilm was formed on metal plates when these were lowered in a beaker with white water. Three different beakers were set up, one with addition of a biocide with active component 4,5-DICHLORO-1,2-DITHIOLONE from the start, one with the addition of the same biocide after three days and one with no addition at all of biocide. Samples were taken from the beakers and analyzed with T-RFLP.

     

    In this report, we show that biocides affect planktonic and biofilm micro-organisms differently. There are however some micro-organisms in the biofilm that does not get affected by the biocide.

     

    The experimental in this report is a good way of investigate the influence that biocides have on planktonic and biofilm micro-organisms, but to get even greater result the experiment should be done over a longer period of time and repeatedly.

  • 57.
    Bourghardt, Johan
    et al.
    Wallenberg Laboratory for Cardiovascular Research, Göteborg University, Sahlgrenska University Hospital, Göteborg, Sweden.
    Bergström, Göran
    Wallenberg Laboratory for Cardiovascular Research, Göteborg University, Sahlgrenska University Hospital, Göteborg, Sweden.
    Krettek, Alexandra
    Wallenberg Laboratory for Cardiovascular Research, Göteborg University, Sahlgrenska University Hospital, Göteborg, Sweden.
    Sjöberg, Sara
    Wallenberg Laboratory for Cardiovascular Research, Göteborg University, Sahlgrenska University Hospital, Göteborg, Sweden.
    Borén, Jan
    Wallenberg Laboratory for Cardiovascular Research, Göteborg University, Sahlgrenska University Hospital, Göteborg, Sweden.
    Tivesten, Åsa
    Wallenberg Laboratory for Cardiovascular Research, Göteborg University, Sahlgrenska University Hospital, Göteborg, Sweden / Wallenberg Laboratory for Cardiovascular Research, Sahlgrenska University Hospital, Göteborg, Sweden.
    The endogenous estradiol metabolite 2-methoxyestradiol reduces atherosclerotic lesion formation in female apolipoprotein E-deficient mice2007In: Endocrinology, ISSN 0013-7227, E-ISSN 1945-7170, Vol. 148, no 9, p. 4128-4132Article in journal (Refereed)
    Abstract [en]

    Estradiol, the major endogenous estrogen, reduces experimental atherosclerosis and metabolizes to 2-methoxyestradiol in vascular cells. Currently undergoing evaluation in clinical cancer trials, 2-methoxyestradiol potently inhibits cell proliferation independently of the classical estrogen receptors. This study examined whether 2-methoxyestradiol affects atherosclerosis development in female mice. Apolipoprotein E-deficient mice, a well-established mouse model of atherosclerosis, were ovariectomized and treated through slow-release pellets with placebo, 17beta-estradiol (6 microg/d), or 2-methoxyestradiol [6.66 microg/d (low-dose) or 66.6 microg/d (high-dose)]. After 90 d, body weight gain decreased and uterine weight increased in the high-dose but not low-dose 2-methoxyestradiol group. En face analysis showed that the fractional area of the aorta covered by atherosclerotic lesions decreased in the high-dose 2-methoxyestradiol (52%) but not in the low-dose 2-methoxyestradiol group. Total serum cholesterol levels decreased in the high- and low-dose 2-methoxyestradiol groups (19%, P < 0.05 and 21%, P = 0.062, respectively). Estradiol treatment reduced the fractional atherosclerotic lesion area (85%) and decreased cholesterol levels (42%). In conclusion, our study shows for the first time that 2-methoxyestradiol reduces atherosclerotic lesion formation in vivo. The antiatherogenic activity of an estradiol metabolite lacking estrogen receptor activating capacity may argue that trials on cardiovascular effects of hormone replacement therapy should use estradiol rather than other estrogens. Future research should define the role of 2-methoxyestradiol as a mediator of the antiatherosclerotic actions of estradiol. Furthermore, evaluation of the effects of 2-methoxyestradiol on cardiovascular disease endpoints in ongoing clinical trials is of great interest.

  • 58.
    Bourghardt, Johan
    et al.
    Wallenberg Laboratory for Cardiovascular Research, Sahlgrenska University Hospital, University of Gothenburg, Gothenburg, Sweden.
    Wilhelmson, Anna S. K.
    Wallenberg Laboratory for Cardiovascular Research, Sahlgrenska University Hospital, University of Gothenburg, Gothenburg, Sweden.
    Alexanderson, Camilla
    Wallenberg Laboratory for Cardiovascular Research, Sahlgrenska University Hospital, University of Gothenburg, Gothenburg, Sweden.
    De Gendt, Karel
    Laboratory for Experimental Medicine and Endocrinology, Department of Experimental Medicine, Katholieke Universiteit Leuven, Leuven, Belgium.
    Verhoeven, Guido
    Laboratory for Experimental Medicine and Endocrinology, Department of Experimental Medicine, Katholieke Universiteit Leuven, Leuven, Belgium.
    Krettek, Alexandra
    Nordic School of Public Health NHV, Gothenburg, Sweden.
    Ohlsson, Claes
    Center for Bone Research, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
    Tivesten, Åsa
    Wallenberg Laboratory for Cardiovascular Research, Sahlgrenska University Hospital, University of Gothenburg, Gothenburg, Sweden.
    Androgen receptor-dependent and independent atheroprotection by testosterone in male mice2010In: Endocrinology, ISSN 0013-7227, E-ISSN 1945-7170, Vol. 151, no 11, p. 5428-5437Article in journal (Refereed)
    Abstract [en]

    The atheroprotective effect of testosterone is thought to require aromatization of testosterone to estradiol, but no study has adequately addressed the role of the androgen receptor (AR), the major pathway for the physiological effects of testosterone. We used AR knockout (ARKO) mice on apolipoprotein E-deficient background to study the role of the AR in testosterone atheroprotection in male mice. Because ARKO mice are testosterone deficient, we sham operated or orchiectomized (Orx) the mice before puberty, and Orx mice were supplemented with placebo or a physiological testosterone dose. From 8 to 16 wk of age, the mice consumed a high-fat diet. In the aortic root, ARKO mice showed increased atherosclerotic lesion area (+80%, P < 0.05). Compared with placebo, testosterone reduced lesion area both in Orx wild-type (WT) mice (by 50%, P < 0.001) and ARKO mice (by 24%, P < 0.05). However, lesion area was larger in testosterone-supplemented ARKO compared with testosterone-supplemented WT mice (+57%, P < 0.05). In WT mice, testosterone reduced the presence of a necrotic core in the plaque (80% among placebo-treated vs. 12% among testosterone-treated mice; P < 0.05), whereas there was no significant effect in ARKO mice (P = 0.20). In conclusion, ARKO mice on apolipoprotein E-deficient background display accelerated atherosclerosis. Testosterone treatment reduced atherosclerosis in both WT and ARKO mice. However, the effect on lesion area and complexity was more pronounced in WT than in ARKO mice, and lesion area was larger in ARKO mice even after testosterone supplementation. These results are consistent with an AR-dependent as well as an AR-independent component of testosterone atheroprotection in male mice.

  • 59. Bradley, Jean-Claude
    et al.
    Guha, Rajarshi
    Lang, Andrew
    Lindenbaum, Pierre
    Neylon, Cameron
    Williams, Antony
    Willighagen, Egon
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Beautifying Data in the Real World2009In: Beautiful Data: The Stories Behind Elegant Data Solutions / [ed] Toby Segaran & Jeff Hammerbacher, Sebastol, USA: O'Reilly , 2009, 1, p. 259-278Chapter in book (Other (popular science, discussion, etc.))
  • 60.
    Broman, T
    et al.
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Thelaus, J
    Andersson, A-C
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Bäckman, S
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Wikström, P
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Larsson, E
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Granberg, M
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Karlsson, L
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Bäck, E
    Department of Infectious Diseases, Örebro University Hospital, Örebro.
    Eliasson, H
    Department of Infectious Diseases, Örebro University Hospital, Örebro.
    Mattsson, R
    National Veterinary Institute, Uppsala.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Forsman, M
    Department of CBRN Defence and Security, Swedish Defence Research Agency, Umeå.
    Molecular Detection of Persistent Francisella tularensis Subspecies holarctica in Natural Waters2011In: International Journal of Microbiology, ISSN 1687-918X, E-ISSN 1687-9198, Vol. 2011, p. Article ID 851946-Article in journal (Refereed)
    Abstract [en]

    Tularemia, caused by the bacterium Francisella tularensis, where F. tularensis subspecies holarctica has long been the cause of endemic disease in parts of northern Sweden. Despite this, our understanding of the natural life-cycle of the organism is still limited. During three years, we collected surface water samples (n = 341) and sediment samples (n = 245) in two areas in Sweden with endemic tularemia. Real-time PCR screening demonstrated the presence of F. tularenis lpnA sequences in 108 (32%) and 48 (20%) of the samples, respectively. The 16S rRNA sequences from those samples all grouped to the species F. tularensis. Analysis of the FtM19InDel region of lpnA-positive samples from selected sampling points confirmed the presence of F. tularensis subspecies holarctica-specific sequences. These sequences were detected in water sampled during both outbreak and nonoutbreak years. Our results indicate that diverse F. tularensis-like organisms, including F. tularensis subsp. holarctica, persist in natural waters and sediments in the investigated areas with endemic tularemia.

  • 61.
    Brännström, Nikolina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Comparison of the natural variation of Tripeptidyl peptidase II activity in blood samples among healthy subjects2011Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    Tripeptidyl peptidase II (TPPII) is a very large enzyme complex with a molecular weight of 5-6MDa and it has broad substrate specificity. It is located in the cytosol in most eukaryotic cells. TPPII which is an amino peptidase has exopeptidase activity, it removes tripeptides from the free N-terminus of oligopeptides and it is acting downstream of the proteasome. TPPII participates in a number of important processes in the cell: protein degradation, antigen presentation and apoptosis. In some tumor cells an increased expression of TPPII has been found which raise the question if TPPII can be used as a tumor marker in blood. The aim of this study was to compare the natural variation of the enzyme activity in blood samples among healthy subjects and also to see if the specific activity changed dependent on how the samples were stored after sampling. To do this an activity assay was used to measure the TPPII enzyme activity and the method of Bradford. Western blot was used to ensure that the right product, TPPII protein was detected. Finally qPCR was used to evaluate the feasibility of detecting TPPII mRNA in blood samples and to determine if mRNA levels correlated to the TPPII protein amount. The result showed a variation in enzyme activity among healthy subjects, a high activity in erythrocyte fractions compared to plasma and leukocyte fractions and also that storing the samples as lysate in -80C gave the least change in relative specific activity in comparison to the fresh blood cell fractions.

  • 62.
    Buckland, Robert J
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Watt, Danielle L
    Chittoor, Balasubramanyam
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Nilsson, Anna Karin
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Kunkel, Thomas A
    Chabes, Andrei
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics. Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Increased and Imbalanced dNTP Pools Symmetrically Promote Both Leading and Lagging Strand Replication Infidelity2014In: PLOS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 10, no 12, article id e1004846Article in journal (Refereed)
    Abstract [en]

    The fidelity of DNA replication requires an appropriate balance of dNTPs, yet the nascent leading and lagging strands of the nuclear genome are primarily synthesized by replicases that differ in subunit composition, protein partnerships and biochemical properties, including fidelity. These facts pose the question of whether imbalanced dNTP pools differentially influence leading and lagging strand replication fidelity. Here we test this possibility by examining strand-specific replication infidelity driven by a mutation in yeast ribonucleotide reductase, rnr1-Y285A, that leads to elevated dTTP and dCTP concentrations. The results for the CAN1 mutational reporter gene present in opposite orientations in the genome reveal that the rates, and surprisingly even the sequence contexts, of replication errors are remarkably similar for leading and lagging strand synthesis. Moreover, while many mismatches driven by the dNTP pool imbalance are efficiently corrected by mismatch repair, others are repaired less efficiently, especially those in sequence contexts suggesting reduced proofreading due to increased mismatch extension driven by the high dTTP and dCTP concentrations. Thus the two DNA strands of the nuclear genome are at similar risk of mutations resulting from this dNTP pool imbalance, and this risk is not completely suppressed even when both major replication error correction mechanisms are genetically intact.

  • 63.
    Buznyk, Oleksiy
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences. National Academic Medical Science Ukraine, Ukraine.
    Pasyechnikova, Nataliya
    National Academic Medical Science Ukraine, Ukraine.
    Islam, Mohammad Mirazul
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences. Karolinska Institute, Sweden.
    Iakymenko, Stanislav
    National Academic Medical Science Ukraine, Ukraine.
    Fagerholm, Per
    Linköping University, Department of Clinical and Experimental Medicine, Division of Neuro and Inflammation Science. Linköping University, Faculty of Medicine and Health Sciences. Region Östergötland, Anaesthetics, Operations and Specialty Surgery Center, Department of Ophthalmology in Linköping.
    Griffith, May
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Medicine and Health Sciences. Karolinska Institute, Sweden.
    Bioengineered Corneas Grafted as Alternatives to Human Donor Corneas in Three High-Risk Patients2015In: Clinical and Translational Science, ISSN 1752-8054, E-ISSN 1752-8062, Vol. 8, no 5, p. 558-562Article in journal (Refereed)
    Abstract [en]

    Corneas with severe pathologies have a high risk of rejection when conventionally grafted with human donor tissues. In this early observational study, we grafted bioengineered corneal implants made from recombinant human collagen and synthetic phosphorylcholine polymer into three patients for whom donor cornea transplantation carried a high risk of transplant failure. These patients suffered from corneal ulcers and recurrent erosions preoperatively. The implants provided relief from pain and discomfort, restored corneal integrity by promoting endogenous regeneration of corneal tissues, and improved vision in two of three patients. Such implants could in the future be alternatives to donor corneas for high-risk patients, and therefore, merits further testing in a clinical trial.

  • 64.
    Carlsson, Jenny
    et al.
    Linköping University, Department of Physics, Chemistry and Biology, Applied Physics . Linköping University, The Institute of Technology.
    Gullstrand, Camilla
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics . Linköping University, Faculty of Health Sciences.
    Westermark, Gunilla
    Linköping University, Department of Clinical and Experimental Medicine, Cell Biology. Linköping University, Faculty of Health Sciences.
    Ludvigsson, Johnny
    Linköping University, Department of Clinical and Experimental Medicine, Pediatrics . Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre of Paediatrics and Gynecology and Obstetrics, Department of Paediatrics in Linköping.
    Enander, Karin
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics . Linköping University, The Institute of Technology.
    Liedberg, Bo
    Linköping University, Department of Physics, Chemistry and Biology, Sensor Science and Molecular Physics . Linköping University, The Institute of Technology.
    An indirect competitive immunoassay for insulin autoantibodies based on surface plasmon resonance2008In: Biosensors and Bioelectronics, ISSN 0956-5663, Vol. 24, no 4, p. 876-881Article in journal (Refereed)
    Abstract [en]

    We have developed a sensitive and specific method based on surface plasmon resonance (SPR) for detection of insulin autoantibodies (IAA) in serum samples from individuals at high risk of developing type 1 diabetes (T1D). When measuring trace molecules in undiluted sera with label-free techniques like SPR, non-specific adsorption of matrix proteins to the sensor surface is often a problem, since it causes a signal that masks the analyte response. The developed method is an indirect competitive immunoassay designed to overcome these problems. Today, IAA is mainly measured in radio immunoassays (RIAs), which are time consuming and require radioactively labeled antigen. With our SPR-based immunoassay the overall assay time is reduced by a factor of >100 (4 days to 50 min), while sensitivity is maintained at a level comparable to that offered by RIA.

  • 65.
    Carlsson, Lena
    et al.
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy.
    Fischer, Christine
    Karoloinska Institute.
    Sjöberg, Gunnnar
    Karoloinska Institute.
    Robson, Richard M
    Iowa Statte University.
    Sejersen, Thomas
    Karoloinska Institute.
    Thornell, Lars-Eric
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy.
    Cytoskeletal derangements in hereditary myopathy with a desmin L345P mutation2002In: Acta Neuropathologica, ISSN 0001-6322, E-ISSN 1432-0533, Vol. 104, no 5, p. 493-504Article in journal (Refereed)
    Abstract [en]

    Patients with abnormal accumulations of desmin have been described in myopathies with or without cardiac involvement. Desmin deposits were sometimes associated with abnormal aggregates of other cytoskeletal proteins. In the present study we present how the cytoskeletal organisation of desmin, nestin, synemin, paranemin, plectin and alphaB-crystallin is altered in skeletal muscles from a patient with a L345P mutation in the desmin gene. In general, accumulations of desmin together with synemin, nestin, plectin and alphaB-crystallin were present between myofibrils and beneath the sarcolemma. However, as the biopsy samples were very myopathic, large variability in fibre size and fibre maturation was seen, thus the myofibrillar content and the cytoskeletal organisation varied considerably. In cultured satellite cells from the patient, desmin aggregates were not observed in initial passages, but occurred over time in culture in the form of perinuclear, peripheral or cytoplasmic deposits. Nestin colocalised to the abnormal desmin deposits to a larger extent than did vimentin. alphaB-Crystallin was only present in cells with a disrupted desmin network. Plectin was altered in a subset of cells with a disrupted desmin network, whereas synemin and paranemin were not detected. We conclude that the L345P desmin mutation has a profound influence on the cytoskeletal organisation both in vivo and in vitro, which reflects the pathogenesis of the desmin myopathy.

  • 66.
    Carlsson, Lena
    et al.
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy.
    Li, Z L
    Université Paris.
    Paulin, D
    Université Paris.
    Price, M G
    Baylor College of Medicine.
    Breckler, J
    San Fransisco State University.
    Robson, R M
    Iowa State University.
    Wiche, G
    University of Vienna.
    Thornell, Lars-Eric
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy.
    Differences in the distribution of synemin, paranemin, and plectin in skeletal muscles of wild-type and desmin knock-out mice2000In: Histochemistry and Cell Biology, ISSN 0948-6143, E-ISSN 1432-119X, Vol. 114, no 1, p. 39-47Article in journal (Refereed)
    Abstract [en]

    Mice lacking the gene encoding for the intermediate filament protein desmin have a surprisingly normal myofibrillar organization in skeletal muscle fibers, although myopathy develops in highly used muscles. In the present study we examined how synemin, paranemin, and plectin, three key cytoskeletal proteins related to desmin, are organized in normal and desmin knock-out (K/O) mice. We show that in wild-type mice, synemin, paranemin, and plectin were colocalized with desmin in Z-disc-associated striations and at the sarcolemma. All three proteins were also present at the myotendinous junctions and in the postsynaptic area of motor endplates. In the desmin K/O mice the distribution of plectin was unaffected, whereas synemin and paranemin were partly affected. The Z-disc-associated striations were in general no longer present in between the myofibrils. In contrast, at the myotendinous and neuromuscular junctions synemin and paranemin were still present. Our study shows that plectin differs from synemin and paranemin in its binding properties to the myofibrillar Z-discs and that the cytoskeleton in junctional areas is particularly complex in its organization.

  • 67.
    Carlsson, Lena
    et al.
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy.
    Thornell, Lars-Eric
    Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB), Anatomy.
    Desmin-related myopathies in mice and man2001In: Acta Physiologica Scandinavica, ISSN 0001-6772, E-ISSN 1365-201X, Vol. 171, no 3, p. 341-8Article in journal (Refereed)
    Abstract [en]

    Desmin, the main intermediate filament (IF) protein in skeletal and heart muscle cells, is of great importance as a part of the cytoskeleton. The IFs surround and interlink myofibrils, and connect the peripheral myofibrils with the sarcolemma. In myotendinous junctions and neuromuscular junctions of skeletal muscle fibres, desmin is enriched. In the heart, desmin is increased at intercalated discs, the attachment between cardiomyocytes, and it is the main component in Purkinje fibres of the conduction system. Desmin is the first muscle-specific protein to appear during myogenesis. Nevertheless, lack of desmin, as shown from experiments with desmin knockout (K/O) mice, does not influence myogenesis or myofibrillogenesis. However, the desmin knock-out mice postnatally develop a cardiomyopathy and a muscle dystrophy in highly used skeletal muscles. In other skeletal muscles the organization of myofibrils is remarkably unaffected. Thus, the main consequence of the lack of desmin is that the muscle fibres become more susceptible to damage. The loss of membrane integrity leads to a dystrophic process, with degeneration and fibrosis. In the heart cardiac failure develops, whereas in affected skeletal muscles regenerative attempts are seen. In humans, accumulations of desmin have been a hallmark for presumptive desmin myopathies. Recent investigations have shown that some families with such a myopathy have a defect in the gene coding for alphaB-crystallin, whereas others have mutations in the desmin gene. Typical features of these patients are cardiac affections and muscle weakness. Thus, mutations in the desmin gene is pathogenic for a distinct type of muscle disorder.

  • 68.
    Chaabane, Wiem
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences. Tunis University, Tunisia.
    Cieślar-Pobuda, Artur
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences. Silesian University of Technology, Gliwice, Poland.
    El-Gazzah, Mohamed
    Tunis University, Tunisia.
    Jain, Mayur V.
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences.
    Rzeszowska-Wolny, Joanna
    Silesian University of Technology, Gliwice, Poland.
    Rafat, Mehrdad
    Linköping University, Department of Biomedical Engineering, Biomedical Instrumentation. Linköping University, Faculty of Health Sciences.
    Stetefeld, Joerg
    University of Manitoba, Winnipeg, Canada.
    Ghavami, Saeid
    University of Manitoba, Winnipeg, Canada.
    Los, Marek
    Linköping University, Department of Clinical and Experimental Medicine, Division of Cell Biology. Linköping University, Faculty of Health Sciences. Pomeranian Medical University, Szczecin, Poland.
    Human-Gyrovirus-Apoptin Triggers Mitochondrial Death Pathway—Nur77 is Required for Apoptosis Triggering: 2014In: Neoplasia, ISSN 1522-8002, E-ISSN 1476-5586, Vol. 16, no 9, p. 679-693Article in journal (Refereed)
    Abstract [en]

    The human gyrovirus derived protein Apoptin (HGV-Apoptin) a homologue of the chicken anemia virus Apoptin (CAV-Apoptin), a protein with high cancer cells selective toxicity, trigger apoptosis selectively in cancer cells. In this paper, we show that HGV-Apoptin acts independently from the death receptor pathway as it induces apoptosis in similar rates in Jurkat cells deficient in either FADD-function or caspase-8 (key players of the extrinsic pathway) and their parental clones. HGV-Apoptin induces apoptosis via the activation of the mitochondrial intrinsic pathway. It induces both mitochondrial inner and outer membrane permebilization, characterized by the loss of the mitochondrial potential and the release into cytoplasm of the pro-apoptotic molecules including apoptosis inducing factor (AIF) and cytochrome c. HGV-Apoptin acts via the apoptosome, as lack of expression of APAF1 in murine embryonic fibroblast strongly protected the cells from HGV-Apoptin-induced apoptosis. Moreover, QVD-oph a broad-spectrum caspase inhibitor delayed HGV-Apoptin-induced death. On the other hand, overexpression of the anti-apoptotic BCL-XL confers resistance to HGV-Apoptin induced cell death. In contrast, cells that lack the expression of the pro-apoptotic BAX and BAK are protected from HGV-Apoptin induced apoptosis. Furthermore, HGV-Apoptin acts independently from p53 signal but triggers the cytoplasmic translocation of Nur77. Taking together this data indicate that HGV-Apoptin acts through the mitochondrial pathway, in a caspase-dependent manner but independently from the death receptor pathway.

  • 69.
    Chi, Celestine N.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Bach, Anders
    University of Copenhagen.
    Gottschalk, Marie
    University of Copenhagen.
    Kristensen, S. Anders
    University of Copenhagen.
    Strømgaard, Kristian
    University of Copenhagen.
    Jemth, Per
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Deciphering the kinetic binding mechanism of dimeric ligands, using a potent plasma-stable dimeric inhibitor of postsynaptic density protein-95 as an example2010In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 285, no 36, p. 28252-28260Article in journal (Refereed)
    Abstract [en]

    Dimeric ligands can be potent inhibitors of protein-protein or enzyme-substrate interactions. They have increased affinity and specificity towards their targets due to their ability to bind simultaneously to two binding sites and are therefore very attractive in drug design. However, few studies have addressed the kinetic mechanism of interaction of such bivalent ligands. We have investigated the binding interaction of a recently identified potent plasma-stable dimeric pentapeptide of PDZ1-2 of PSD-95 using protein engineering in combination with fluorescence polarisation, isothermal titration calorimetry and stopped-flow fluorimetry. Our experiments demonstrate that binding occurs via a two-step process, where an initial binding to either one of the two PDZ domains is followed by an intramolecular step, which produces the bidentate complex. We have determined all rate constants involved in the binding reaction and we also find evidence for a conformational transition of the complex. Our data demonstrate the importance of a slow dissociation for a successful dimeric ligand, but also highlight the possibility of optimizing the intramolecular association rate. The results may therefore aid the design of dimeric inhibitors in general.

  • 70.
    Chorell, Elin
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Mapping the consequenses of physical exercise and nutrition on human health: A predictive metabolomics approach2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Human health is a complex and wide-ranging subject far beyond nutrition and physical exercise. Still, these factors have a huge impact on global health by their ability to prevent diseases and thus promote health. Thus, to identify health risks and benefits, it is necessary to reveal the underlying mechanisms of nutrition and exercise, which in many cases follows a complex chain of events. As a consequence, current health research is generating massive amounts of data from anthropometric parameters, genes, proteins, small molecules (metabolites) et cetera, with the intent to understand these mechanisms. For the study of health responses, especially related to physical exercise and nutrition, alterations in small molecules (metabolites) are in most cases immediate and located close to the phenotypic level and could therefore provide early signs of metabolic imbalances. Since there are roughly as many different responses to exercise and nutrients as there are humans, this quest is highly multifaceted and will benefit from an interpretation of treatment effects on a general as well as on an individual level. This thesis involves the application of chemometric methods to the study of global metabolic reactions, i.e. metabolomics, in a strategy coined predictive metabolomics. Via the application of predictive metabolomics an extensive hypothesis-free biological interpretation has been carried out of metabolite patterns in blood, acquired using gas chromatography-mass spectrometry (GC-MS), related to physical exercise, nutrition and diet, all in the context of human health. In addition, the chemometrics methodology have computational benefits concerning the extraction of relevant information from information-rich data as well as for interpreting general treatment effects and individual responses, as exemplified throughout this work. Health concerns all lifestages, thus this thesis presents a strategic framework in combination with comprehensive interpretations of metabolite patterns throughout life. This includes a broad range of human studies revealing metabolic patterns related to the impact of physical exercise, macronutrient modulation and different fitness status in young healthy males, short and long term dietary treatments in overweight post menopausal women as well as metabolic responses related to probiotics treatment and early development in infants. As a result, the studies included in the thesis have revealed metabolic patterns potentially indicative of an anti-catabolic response to macronutrients in the early recovery phase following exercise. Moreover, moderate differences in the metabolome associated with cardiorespiratory fitness level were detected, which could be linked to variation in the inflammatory and antioxidaive defense system. This work also highlighted mechanistic information that could be connected to dietary related weight loss in overweight and obese postmenopausal women in relation to short as well as long term dietary effects based on different macronutrient compositions. Finally, alterations were observed in metabolic profiles in relation to probiotics treatment in the second half of infancy, suggesting possible health benefits of probiotics supplementation at an early age.

     

  • 71. Chung, L. W.
    et al.
    Hayashi, S.
    Lundberg, Marcus
    Kyoto University.
    Nakatsu, T.
    Kato, H.
    Morokuma, K.
    Mechanism of efficient firefly bioluminescence via adiabatic transition state and seam of sloped conical intersection.2008In: Journal of the American Chemical Society, ISSN 0002-7863, E-ISSN 1520-5126, Vol. 130, no 39, p. 12880-12881Article in journal (Refereed)
    Abstract [en]

    Firefly emission is a well-known efficient bioluminescence. However, the mystery of the efficient thermal generation of electronic excited states in firefly still remains unsolved, particularly at the atomic and molecular levels. We performed SA-CASSCF(12,12)/6-31G* and CASPT2(12,12)/6-31G*//SA-CASSCF(12,12)/6-31G* calculations to elucidate the reaction mechanism of bioluminescence from the firefly dioxetanone in the gas phase. Adiabatic transition state (TS) for the O-O bond cleavage and the minimum energy conical intersection (MECI) were located and characterized. The unique topology of MECI featuring a seam of a sloped conical intersection for the firefly dioxetanone, which was uncovered for the first time, emerges along the reaction pathway to provide a widely extended channel to diabatically access the excited-state from the ground state.

  • 72.
    Co, Michelle
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Pressurised Fluid Extraction of Bioactive Species in Tree Barks: Analysis using Hyphenated Electrochemical Mass Spectrometric Detection2010Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Analytical chemistry has developed throughout time to meet current needs. At present, the interest in biorefinery is growing, due to environmental awareness and the depletion of fossil resources. Biomass from agricultural and forestry industries has proven to be excellent raw material for different processes. Biorefinering valuable species such as bioactive species from biomass, without compromising the primary process of the biomass is highly desirable. Pressurised fluid extraction (PFE) using water and ethanol as a solvent was developed for extracting betulin from birch (Betula pendula) bark. Apart from betulin, stilbene glucosides such as astringin, isorhapontin and picied were also extracted from spruce (Picea abies) using PFE. PFE is an advanced technique that extracts at temperatures above the solvent’s atmospheric boiling point. The applied pressure in PFE is mainly to maintain the liquid state of the extraction solvent. Parameters such as type of solvent, temperature, and time affect the extraction selectivity and efficiency. Therefore it is necessary to comprehend these parameters in order to optimise extraction. The DPPH (1,1-diphenyl-2-picrylhydrazyl) assay was used to determine the antioxidant capacity and activity of the obtained bioactive species. The results showed high antioxidant capacity in bioactive species that were extracted at an elevated temperature, 180°C. Extraction and degradation occur simultaneously during the extraction. Hence, it is crucial to separate these two processes in order to obtain the actual value.

    An online hyphenated system of chromatographic separation electrochemical mass spectrometric detection was developed (LC-DAD-ECD-MS/MS). The electrochemical detector facilitates real-time monitoring of the antioxidant capacity and activity of each antioxidant and its oxidation products. This developed LC-DAD-ECD-MS/MS method enabled rapid screening of antioxidants and created a fingerprint map for their oxidation products. Characterisation and molecular elucidation of bioactive species were also performed. Degradation of bioactive species was investigated with the said online system and birch bark extract was compared with birch bark extracts that were hydrothermally treated. The obtained results showed some degradation of antioxidants at 180°C.

    In summary, the aim of this thesis was to develop analytical methods integrated with sustainable chemistry for extraction of bioactive species in biomass from the forestry industry. A novel online system using selective and sensitive detectors such as diode-array, electrochemical, and tandem mass spectrometry was developed to rapidly determine the antioxidant capacity and activity of antioxidants. Furthermore, tandem mass spectrometry enables identification of unknown bioactive species without the need of reference samples.

  • 73.
    Co, Michelle
    et al.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Fagerlund, Amelie
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry.
    Engman, Lars
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry - BMC.
    Sunnerheim, Kerstin
    Department of Natural Sciences, Engineering and Mathematics, Mid Sweden University.
    Sjöberg, Per J. R
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Physical and Analytical Chemistry, Analytical Chemistry.
    Turner, Charlotta
    Dept of Organic Chemistry, Lund University.
    Extraction of Antioxidants from Spruce (Picea abies) Bark using Eco-Friendly Solvents2012In: Phytochemical Analysis, ISSN 0958-0344, E-ISSN 1099-1565, Vol. 23, no 1, p. 1-11Article in journal (Refereed)
    Abstract [en]

    Introduction-Antioxidants are known to avert oxidation processes and they are found in trees and other plant materials. Tree bark is a major waste product from paper pulp industries; hence it is worthwhile to develop an extraction technique to extract the antioxidants.

    Objective- To develop a fast and environmentally sustainable extraction technique for the extraction of antioxidants from bark of spruce (Picea abies) and also to identify the extracted antioxidants that are abundant in spruce bark.

    Methodology- A screening experiment that involved three different techniques, was conducted to determine the best technique to extract antioxidants.The antioxidant capacity of the extracts was determined with DPPH (2,2-diphenyl-2’-picrylhydrazyl) assay. Pressurised fluid extraction (PFE) turned out to be the best technique and a response surface design was therefore utilised to optimise PFE. Furthermore, NMR and HPLC-DAD-MS/MS were applied to identify the extracted antioxidants.

    Results- PFE using water and ethanol as solvent at 160 and 180°C, respectively, gave extracts of the highest antioxidant capacity. Stilbene glucosides such as isorhapontin, piceid and astringin were identified in the extracts.

    Conclusion-The study has shown that PFE is a fast and environmentally sustainable technique, using water and ethanol as solvent for the extraction of antioxidants from spruce bark.

  • 74.
    Colnerud Nilsson, Emma
    Linköping University, Department of Physics, Chemistry and Biology.
    Database for targeted drug screening with Liquid Chromatography - Time-Of-Flight Mass Spectrometry, (LC-TOFMS)2010Independent thesis Advanced level (degree of Master (One Year)), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    Today there are no fully general analytical techniques available for detection and confirmation of known and unknown substances in toxicological screening, further tools are therefore needed. The development of mass spectrometry with time-of-flight (TOF) detection is promising but there are still areas to be further developed and evaluated, both instrumentation and applications.

    During 2009 The National Board of Forensic Medicine-Department of Forensic Genetics and Forensic Toxicology, (RMV) started cooperation with the instrumentation company Waters (Manchester, UK) and the Department of Clinical Pharmacology (KI, Solna) evaluating a new TOF-instrument for toxicological screening. My assignment as a part of this project has been to create a limited and relevant database of drugs and toxics in Excel, including monoisotopic mass, used when screening for pharmaceutical substances and their metabolites most probable to be found in Swedish autopsy material.

    A limited database has been developed based on information from several sources, it ended up in 875 analytes and metabolites. A limited but complete database is more reliable in practise than a big database, by means of a lower frequency of isobars and more information included (e.g. retention time from liquid chromatography) making analysis faster. Commercial databases are generally theoretical, lacking information about for example retention time that often is an important criterion for identification.

  • 75. Cox, Nicholas
    et al.
    Ho, Felix M.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science.
    Pewnim, Naray
    Steffen, Ronald
    Smith, Paul J.
    Havelius, Kajsa G. V.
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science.
    Hughes, Joseph L.
    Debono, Lesley
    Styring, Stenbjörn
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Photochemistry and Molecular Science, Molecular Biomimetics.
    Krausz, Elmars
    Pace, Ron J.
    The S-1 split signal of photosystem II: a tyrosine-manganese coupled interaction2009In: Biochimica et Biophysica Acta - Bioenergetics, ISSN 0005-2728, E-ISSN 1879-2650, Vol. 1787, no 7, p. 882-889Article in journal (Refereed)
    Abstract [en]

    Detailed optical and EPR analyses of states induced in dark-adapted PS II membranes by cryogenic illumination permit characterization and quantification of all pigment derived donors and acceptors, as well as optically silent (in the visible, near infrared) species which are EPR active. Near complete turnover formation of Q(A)(-) is seen in all centers, but with variable efficiency, depending on the donor species. In minimally detergent-exposed PS II membranes, negligible (<5%) oxidation of chlorophyll or carotenoid centers occurs for illumination temperatures 5-20 K. An optically silent electron donor to P680(+) is observed with the same decay kinetics as the S-1 split signal. Cryogenic donors to P680(+) seen are: (i) transient (t(1/2)similar to 150 s) tyrosine related species, including 'split signals' (similar to 15% total centers), (ii) reduced cytochrome b(559) (similar to 30-50% centers), and (iii) an organic donor, possibly an amino acid side chain, (similar to 30% centers).

  • 76. Cribiù, Riccardo
    et al.
    Borbas, K. Eszter
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Cumpstey, Ian
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    On the synthesis of vinyl and phenyl C-furanosides by stereospecific debenzylative cycloetherification2009In: Tetrahedron, ISSN 0040-4020, E-ISSN 1464-5416, Vol. 65, no 10, p. 2022-2031Article in journal (Refereed)
  • 77. Crona, Mikael
    et al.
    Hofer, Anders
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Astorga-Wells, Juan
    Sjöberg, Britt-Marie
    Tholander, Fredrik
    Biochemical Characterization of the Split Class II Ribonucleotide Reductase from Pseudomonas aeruginosa2015In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, no 7, article id e0134293Article in journal (Refereed)
    Abstract [en]

    The opportunistic pathogen Pseudomonas aeruginosa can grow under both aerobic and anaerobic conditions. Its flexibility with respect to oxygen load is reflected by the fact that its genome encodes all three existing classes of ribonucleotides reductase (RNR): the oxygen-dependent class I RNR, the oxygen-indifferent class II RNR, and the oxygen-sensitive class III RNR. The P. aeruginosa class II RNR is expressed as two separate polypeptides (NrdJa and NrdJb), a unique example of a split RNR enzyme in a free-living organism. A split class II RNR is also found in a few closely related gamma-Proteobacteria. We have characterized the P. aeruginosa class II RNR and show that both subunits are required for formation of a biologically functional enzyme that can sustain vitamin B12-dependent growth. Binding of the B12 coenzyme as well as substrate and allosteric effectors resides in the NrdJa subunit, whereas the NrdJb subunit mediates efficient reductive dithiol exchange during catalysis. A combination of activity assays and activity-independent methods like surface plasmon resonance and gas phase electrophoretic macromolecule analysis suggests that the enzymatically active form of the enzyme is a (NrdJa-NrdJb) 2 homodimer of heterodimers, and a combination of hydrogen-deuterium exchange experiments and molecular modeling suggests a plausible region in NrdJa that interacts with NrdJb. Our detailed characterization of the split NrdJ from P. aeruginosa provides insight into the biochemical function of a unique enzyme known to have central roles in biofilm formation and anaerobic growth.

  • 78.
    Córdova, Armando
    et al.
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Ibrahem, Ismail
    Casas, Jesús
    Sundén, Henrik
    Engqvist, Magnus
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Reyes, Efraim
    Amino Acid-Catalyzed Neogenesis of Carbohydrates: A Plausible Ancient Transformation2005In: Chemistry - A European Journal, ISSN 0947-6539, E-ISSN 1521-3765, Vol. 11, no 16, p. 4772-4784Article in journal (Refereed)
    Abstract [en]

    Hexose sugars play a fundamental role in vital biochemical processes and their biosynthesis is achieved through enzyme-catalyzed pathways. Herein we disclose the ability of amino acids to catalyze the asymmetric neogenesis of carbohydrates by sequential cross-aldol reactions. The amino acids mediate the asymmetric de novo synthesis of natural L- and D-hexoses and their analogues with excellent stereoselectivity in organic solvents. In some cases, the four new stereocenters are assembled with almost absolute stereocontrol. The unique feature of these results is that, when an amino acid is employed as the catalyst, a single reaction sequence can convert a protected glycol aldehyde into a hexose in one step. For example, proline and its derivatives catalyze the asymmetric neogenesis of allose with >99 % ee in one chemical manipulation. Furthermore, all amino acids tested catalyzed the asymmetric formation of natural sugars under prebiotic conditions, with alanine being the smallest catalyst. The inherent simplicity of this catalytic process suggests that a catalytic prebiotic “gluconeogenesis” may occur, in which amino acids transfer their stereochemical information to sugars. In addition, the amino acid catalyzed stereoselective sequential cross-aldol reactions were performed as a two-step procedure with different aldehydes as acceptors and nucleophiles. The employment of two different amino acids as catalysts for the iterative direct aldol reactions enabled the asymmetric synthesis of deoxysugars with >99 % ee. In addition, the direct amino acid catalyzed C2+C2+C2 methodology is a new entry for the short, highly enantioselective de novo synthesis of carbohydrate derivatives, isotope-labeled sugars, and polyketide natural products. The one-pot asymmetric de novo syntheses of deoxy and polyketide carbohydrates involved a novel dynamic kinetic asymmetric transformation (DYKAT) mediated by an amino acid.

  • 79.
    Dahl, Leif
    Stockholm University, Faculty of Science, Department of Physical, Inorganic and Structural Chemistry.
    Preparation and characterisation of perovskite and noble metal promoted oxide based catalysts1997Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This work concerns efforts to improve or replace noble-metal based catalysts used for car exhaust purification.

    Various oxide based catalysts have been prepared by impregnating washcoats such as (-Al2O3 and SiO2 with appropriate metal ions, and assemblies calcinated at elevated temperatures have been tested for catalytic activity in the oxidation of CO and C3H6 and reduction of NO. SEM-EDS, ATEM-EDS and XRD have been used for phase analysis and for studies of the morphology of catalysts prepared in situ. The perovskite compounds La1-xSrxMe1-2yCuyRuyO3 with Me = Al and Ga have been prepared, in situ and ex situ [SGIW1]respectively, and tested for catalytic activity. Changes in cell parameters and catalytic activity of in-situ LaAl1-xCuxO3 when exposed alternatingly to oxidising and reducing car exhaust atmospheres have been studied. The catalytic activity of various transition metal oxides with and without addition of Pt/Rh or Pd have also been tested.

  • 80.
    Dahlbäcker, Maria
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Utveckling av ett lågkaloripulver med chokladsmak2011Independent thesis Basic level (degree of Bachelor), 10 credits / 15 HE creditsStudent thesis
    Abstract [en]

    Today, obesity is the cause for many diseases such as metabolic syndrome and cardiovascular diseases. Very low calorie diets (VLCD) are formulated foods with 450-800 kilocalories per day intended for use as the sole dietary source of energy during weight loss. These diets have proven useful in short-term weight loss and show improvements in blood pressure, serum lipids and glycaemia in obese people with type 2 diabetes.

    Currently, there is no harmonised legislation throughout the EU for VLCD-products, however Codex Alimentarius has developed a standard (Codex Standard 203-1995) which the Nordic Council of Ministers have based their Nordic recommendations (Report 1993:557) on. The Swedish recommendations are based on this report and The Swedish National Food Administration direct the control and labelling of products merchandised inSweden.

    The current recommendations for VLCD have been summarized in this study. A review of scientific studies of the use of VLCD-products demonstrated the safe use of these products for a limited time to achieve weight loss, due to nutritional composition preventing loss of lean body mass.

    The aim of this study was to formulate a recipe for a chocolate flavoured VLCD-powder based on The Swedish National Food Administration’s VLCD recommendations. First, a theoretical recipe was calculated and based on sensory analysis different sources of raw materials and the amounts were evaluated and adjusted in different trials. In the fifth and final trial, a product with a satisfying taste and texture was created.

    The results from the different trials showed that the low energy content in the product demanded the right type of raw material to achieve a pleasant flavour and correct amount of nutrients. The raw material used in the project was presented and described in the background.

    All together the project gave an overview of VLCD-products whereby no evidence of serious adverse effects was found in the literature from using VLCD for a limited time. In formulating a VLCD powder it was important to use the right raw materials and combination of ingredients to develop a product with a satisfying taste and texture.

     

  • 81.
    Dahlgren, Markus K
    et al.
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Zetterström, Caroline E
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Gylfe, Åsa
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology.
    Linusson, Anna
    Umeå University, Faculty of Science and Technology, Department of Chemistry.
    Elofsson, Mikael
    Umeå University, Faculty of Science and Technology, Department of Chemistry. Umeå University, Faculty of Medicine, Umeå Centre for Microbial Research (UCMR).
    Statistical molecular design of a focused salicylidene acylhydrazide library and multivariate QSAR of inhibition of type III secretion in the Gram-negative bacterium Yersinia2010In: Bioorganic & Medicinal Chemistry, ISSN 0968-0896, E-ISSN 1464-3391, Vol. 18, no 7, p. 2686-2703Article in journal (Refereed)
    Abstract [en]

    A combined application of statistical molecular design (SMD), quantitative structure-activity relationship (QSAR) modeling and prediction of new active compounds was effectively used to develop salicylidene acylhydrazides as inhibitors of type III secretion (T3S) in the Gram-negative pathogen Yersinia pseudotuberculosis. SMD and subsequent synthesis furnished 50 salicylidene acylhydrazides in high purity. Based on data from biological evaluation in T3S linked assays 18 compounds were classified as active and 25 compounds showed a dose-dependent inhibition. The 25 compounds were used to compute two multivariate QSAR models and two multivariate discriminant analysis models were computed from both active and inactive compounds. Three of the models were used to predict 4416 virtual compounds in consensus and eight new compounds were selected as an external test set. Synthesis and biological evaluation of the test set in Y. pseudotuberculosis and the intracellular pathogen Chlamydia trachomatis validated the models. Y. pseudotuberculosis and C. trachomatis cell-based infection models showed that compounds identified in this study are selective and non-toxic inhibitors of T3S dependent virulence.

  • 82.
    Dahlgren, Markus
    et al.
    Umeå University, Faculty of Science and Technology, Chemistry.
    Kauppi, Anna
    Umeå University, Faculty of Science and Technology, Chemistry.
    Olsson, Ing-Marie
    Umeå University, Faculty of Science and Technology, Chemistry.
    Linusson Jonsson, Anna
    Umeå University, Faculty of Science and Technology, Chemistry.
    Elofsson, Mikael
    Umeå University, Faculty of Science and Technology, Chemistry.
    Design, Synthesis, and Multivariate Quantitative Structure-Activity Relationship of Salicylanilides-Potent Inhibitors of Type III Secretion in Yersinia2007In: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 50, no 24, p. 6177-6188Article in journal (Refereed)
    Abstract [en]

    Analogues to the salicylanilide N-(4-Chlorophenyl)-2-acetoxy-3,5-diiodobenzamide, 1a, an inhibitor of type III secretion (T3S) in Yersinia, were selected, synthesized, and biologically evaluated in three cycles. First, a set of analogues with variations in the salicylic acid ring moiety was synthesized to probe possible structural variation. A basic structure-activity relationship was established and then used to cherry-pick compounds from a principal component analysis score plot of salicylanilides to generate a second set. A third set with increased likelihood of biological activity was designed using D-optimal onion design. A quantitative structure-activity relationship model using hierarchical partial least-square regression to latent structures (Hi-PLS) was computed using PLS score vectors of building blocks correlated to the % inhibition of T3S as a response. A PLS discriminant analysis (PLS-DA) model was derived using the same descriptor set as that for the Hi-PLS model. Both models were validated with an external test set.

  • 83.
    Dahlin, Anna M
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Henriksson, Maria L
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Van Guelpen, Bethany
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Stenling, Roger
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Öberg, Åke
    Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
    Rutegård, Jörgen
    Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences, Surgery.
    Palmqvist, Richard
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Colorectal cancer prognosis depends on T-cell infiltration and molecular characteristics of the tumor2011In: Modern Pathology, ISSN 0893-3952, E-ISSN 1530-0285, Vol. 24, p. 671-682Article in journal (Refereed)
    Abstract [en]

    The aim of this study was to relate the density of tumor infiltrating T cells to cancer-specific survival in colorectal cancer, taking into consideration the CpG island methylator phenotype (CIMP) and microsatellite instability (MSI) screening status. The T-cell marker CD3 was stained by immunohistochemistry in 484 archival tumor tissue samples. T-cell density was semiquantitatively estimated and scored 1-4 in the tumor front and center (T cells in stroma), and intraepithelially (T cells infiltrating tumor cell nests). Total CD3 score was calculated as the sum of the three CD3 scores (range 3-12). MSI screening status was assessed by immunohistochemistry. CIMP status was determined by quantitative real-time PCR (MethyLight) using an eight-gene panel. We found that patients whose tumors were highly infiltrated by T cells (total CD3 score ≥7) had longer survival compared with patients with poorly infiltrated tumors (total CD3 score ≤4). This finding was statistically significant in multivariate analyses (multivariate hazard ratio, 0.57; 95% confidence interval, 0.31-1.00). Importantly, the finding was consistent in rectal cancer patients treated with preoperative radiotherapy. Although microsatellite unstable tumor patients are generally considered to have better prognosis, we found no difference in survival between microsatellite unstable and microsatellite stable (MSS) colorectal cancer patients with similar total CD3 scores. Patients with MSS tumors highly infiltrated by T cells had better prognosis compared with intermediately or poorly infiltrated microsatellite unstable tumors (log rank P=0.013). Regarding CIMP status, CIMP-low was associated with particularly poor prognosis in patients with poorly infiltrated tumors (multivariate hazard ratio for CIMP-low versus CIMP-negative, 3.07; 95% confidence interval, 1.53-6.15). However, some subset analyses suffered from low power and are in need of confirmation by independent studies. In conclusion, patients whose tumors are highly infiltrated by T cells have a beneficial prognosis, regardless of MSI, whereas the role of CIMP status in this context is less clear.

  • 84.
    Dahlqvist, Ulla
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Ek, Pia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Engström, Lorentz
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Endogenous substrates of protein kinase in rat liver cell sap under different dietary conditions1978In: Biochimica et Biophysica Acta, ISSN 0006-3002, E-ISSN 1878-2434, Vol. 540, no 1, p. 13-23Article in journal (Refereed)
    Abstract [en]

    Liver cell sap from normally fed rats, rats fed with a high-carbohydrate diet and fasted rats was chromatographed on DEAE-cellulose (pH 7.0). The chromatogram from each diet group was analyzed for pyruvate kinase activity and endogenous substrates of cyclic AMP-stimulated protein kinase. The materials were pooled into five phosphorylatable fractions, in each of which phosphate incorporation at 0.1 mM and 1.0 mM [32P]ATP in the presence of cyclic AMP and protein kinase was determined. For characterization of the phosphorylatable components, thin-layer gel chromatography on Sephadex G-200 and polyacrylamide gel electrophoresis in detergent were used for determination of native and minimal molecular weights, respectively. Except for pyruvate kinase, eight components which incorporated at least 0.05 nmol of [32P]phosphate/g of liver were detected. The phosphorylation of four of them was stimulated by cyclic AMP. Their minimal molecular weights were 42000, 21000, 52000 and 49000. The component with a minimal molecular weight of 42000 seemed to have a native molecular weight of 160000. Both the 21000 and the 52000 component had a native molecular weight of about 110000-120000. The protein with a minimal molecular weight of 49000 could not be correlated with certainty to a native molecular weight. The proteins whose phosphorylation was not stimulated by cyclic AMP had minimal molecular weights of 54000, 39000, 34000 and 22000.

  • 85.
    Danfors, Torsten
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Sörensen, Jens
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Section of Nuclear Medicine and PET.
    Lubberink, Mark
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Section of Medical Physics.
    Kumlien, Eva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Relative Cerbral Blood Flow Measurement using dynamic Flumazenil-PET may Replace Fluorodeoxyglucose-PET in Epilepsy Surgical Investigations2012Article in journal (Other academic)
  • 86.
    David, Lindström
    KTH, School of Chemical Science and Engineering (CHE), Chemistry, Surface and Corrosion Science.
    Galvanized steel in outdoor constructions - metal runoff, corrosion and patina formation2010Licentiate thesis, comprehensive summary (Other academic)
  • 87.
    Degerman, Sofie
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Josefsson, Maria
    Umeå University, Faculty of Social Sciences, Centre for Demographic and Ageing Research (CEDAR).
    Nordin Adolfsson, Annelie
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Psychiatry.
    Wennstedt, Sigrid
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Landfors, Mattias
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Haider, Zahra
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Pudas, Sara
    Umeå University, Faculty of Medicine, Umeå Centre for Functional Brain Imaging (UFBI). Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB).
    Hultdin, Magnus
    Umeå University, Faculty of Medicine, Department of Medical Biosciences.
    Nyberg, Lars
    Umeå University, Faculty of Medicine, Umeå Centre for Functional Brain Imaging (UFBI). Umeå University, Faculty of Medicine, Department of Integrative Medical Biology (IMB). Umeå University, Faculty of Medicine, Department of Radiation Sciences.
    Adolfsson, Rolf
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Psychiatry.
    Maintained memory in aging is associated with young epigenetic age2017In: Neurobiology of Aging, ISSN 0197-4580, E-ISSN 1558-1497, Vol. 55, p. 167-171Article in journal (Refereed)
    Abstract [en]

    Epigenetic alterations during aging have been proposed to contribute to decline in physical and cognitive functions, and accelerated epigenetic aging has been associated with disease and all-cause mortality later in life. In this study, we estimated epigenetic age dynamics in groups with different memory trajectories (maintained high performance, average decline, and accelerated decline) over a 15-year period. Epigenetic (DNA-methylation [DNAm]) age was assessed, and delta age (DNAm age - chronological age) was calculated in blood samples at baseline (age: 55-65 years) and 15 years later in 52 age- and gender-matched individuals from the Betula study in Sweden. A lower delta DNAm age was observed for those with maintained memory functions compared with those with average (p = 0.035) or accelerated decline (p = 0.037). Moreover, separate analyses revealed that DNAm age at follow-up, but not chronologic age, was a significant predictor of dementia (p = 0.019). Our findings suggest that young epigenetic age contributes to maintained memory in aging.

  • 88.
    Diesen, Jarle Sidney
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Chemistry, Organic Chemistry.
    Asymmetric Hydrogenations of Imines, Vinyl Fluorides, Enol Phosphinates and Other Alkenes Using N,P-Ligated Iridium Complexes2008Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The research described in this thesis is directed toward the efficient, enantioselective synthesis of chiral products that have useful functionality. This goal was pursued through catalytic asymmetric hydrogenation, a reaction class that selectively introduces one or two stereocenters into a molecule in an atom-efficient step. This reaction uses a small amount (often <1 mol%) of a chiral catalyst to impart stereoselectivity to the product formed. Though catalytic asymmetric hydrogenation is not a new reaction type, there remain many substrate classes for which it is ineffective. The present thesis describes efforts to extend the reaction to some of these substrates classes. Some of the products synthesized in these studies may eventually find use as building blocks for the production of chiral pharmaceuticals, agrochemicals, or flavouring or colouring agents. However, the primary and immediate aim of this thesis was to develop and demonstrate new catalysts that are rapid and effective in the asymmetric hydrogenation of a broad range of compounds.

    Paper I describes the design and construction of two new, related chiral iridium compounds that are catalysts for asymmetric hydrogenation. They each contain an N,P-donating phosphinooxazoline ligand that is held together by a rigid bicyclic unit. One of these iridium compounds catalyzed the asymmetric hydrogenation of acyclic aryl imines, often with very good enantioselectivities. This is particularly notable because acyclic imines are difficult to reduce with useful enantioselectivity. The second catalyst was useful for the asymmetric hydrogenation of two aryl olefins. In Paper II, the class of catalysts introduced into Paper I is expanded to include many more related compounds, and these are also applied to the asymmetric hydrogenation of prochiral imines and olefins. By studying a range of related catalysts that differ in a single attribute, we were able to probe how different parts of the catalyst affect the yield and selectivity of the hydrogenation reactions.

    Whereas iridium catalysts had been applied to the asymmetric hydrogenation of imines and largely unfunctionalized olefins prior to this work (with varied degrees of success), they had not been used to reduce fluoroolefins. Their hydrogenation, which is discussed in Paper III, was complicated by concomitant defluorination to yield non-halogenated alkanes. To combat this problem, several iridium-based hydrogenation catalysts were applied to the reaction. Two catalysts stood out for their ability to produce chiral fluoroalkanes in good enantioselectivity while minimizing the defluorination reaction, and one of these bore a phosphinooxazoline ligand of the type described in Papers I and II.

    Enol phosphinates are another class of olefins that had not previously been subjected to iridium-catalyzed asymmetric hydrogenation. They do, however, constitute an attractive substrate class, because the product chiral alkyl phosphinates can be transformed into chiral alcohols or chiral phosphines with no erosion of enantiopurity. Iridium complexes of the phosphinooxazoline ligands described in Papers I and II were extremely effective catalysts for the asymmetric hydrogenation of enol phosphinates. They produced alkyl phosphinates from di- and trisubstituted enol phosphinate, β-ketoester-derived enol phosphinates, and even purely alkyl-substituted enol phopshinates, in very high yields and enantioselectivities.

  • 89.
    Dinér, Peter
    Uppsala University, Disciplinary Domain of Science and Technology, Chemistry, Department of Biochemistry and Organic Chemistry, Organic Chemistry.
    Catalytic asymmetric chiral lithium amide-promoted epoxide rearrangement: a NMR spectroscopic and kinetic investigation2010In: Tetrahedron: asymmetry, ISSN 0957-4166, E-ISSN 1362-511X, Vol. 21, no 21-22, p. 2733-2739Article in journal (Refereed)
    Abstract [en]

    The lithium amide derived from the chiral diamine (1R,3S,4S)-3-(1-pyrrolidinyl)methyl-2-azabicyclo[2.2.1]heptane, has been reported to catalytically deprotonate cyclohexene oxide and other epoxides, yielding chiral allylic alcohols in excellent enantiomeric excess. In this work, 6Li, 1H and 13 C NMR spectroscopy have been used to study the aggregation of the chiral lithium amide in THF and the influence on the aggregation by the addition of additives, such as 1,8-diazabicyclo-[5.4.0]undec-7-ene (DBU). The activated complex under catalytic deprotonation of cyclohexene oxide, that is, with excess Li-DBU and free DBU, is built from one monomer of the chiral lithium amide, one molecule of epoxide and one additional molecule of DBU. The reaction order (0.97) obtained for the bulk base Li-DBU shows an inverse dependence on the concentration, suggesting a deaggregation of the initial mixed dimer to a monomer-based transition state containing a monomer of the lithium amide.

  • 90.
    Dollery, Clare M.
    et al.
    Leducq Ctr. for Cardiovasc. Research, Department of Medicine, Brigham and Women's Hospital, Boston, MA, United States.
    Owen, Caroline A.
    Cardiovascular Division, Department of Medicine, Brigham and Women's Hospital, Boston, MA, United States.
    Sukhova, Galina K
    Leducq Ctr. for Cardiovasc. Research, Department of Medicine, Brigham and Women's Hospital, Boston, MA, United States.
    Krettek, Alexandra
    Leducq Ctr. for Cardiovasc. Research, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, United States.
    Shapiro, Steven D.
    Cardiovascular Division, Department of Medicine, Brigham and Women's Hospital, Boston, MA, United States.
    Libby, Peter
    Leducq Ctr. for Cardiovasc. Research, Department of Medicine, Brigham and Women's Hospital, Boston, MA, United States / Brigham and Women's Hospital, Harvard Medical School, EBRC 307, 221 Longwood Ave., Boston, MA 02115, United States.
    Neutrophil elastase in human atherosclerotic plaques: production by macrophages2003In: Circulation, ISSN 0009-7322, E-ISSN 1524-4539, Vol. 107, no 22, p. 2829-2836Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Catabolism of the extracellular matrix (ECM) contributes to vascular remodeling in health and disease. Although metalloenzymes and cysteinyl proteinases have garnered much attention in this regard, the role of serine-dependent proteinases in vascular ECM degradation during atherogenesis remains unknown. We recently discovered the presence of the metalloproteinase MMP-8, traditionally associated only with neutrophils, in atheroma-related cells. Human neutrophil elastase (NE) plays a critical role in lung disease, but the paucity of neutrophils in the atheromatous plaque has led to neglect of its potential role in vascular biology. NE can digest elastin, fibrillar and nonfibrillar collagens, and other ECM components in addition to its ability to modify lipoproteins and modulate cytokine and MMP activity.

    METHODS AND RESULTS: Fibrous and atheromatous plaques but not normal arteries contained NE. In particular, NE abounded in the macrophage-rich shoulders of atheromatous plaques with histological features of vulnerability. Neutrophil elastase and macrophages colocalized in such vulnerable plaques (n=7). In situ hybridization revealed NE mRNA in macrophage-rich areas, indicating local production of this enzyme. Freshly isolated blood monocytes, monocyte-derived macrophages, and vascular endothelial cells in culture produced active NE and contained NE mRNA. Monocytes produced NE constitutively, with little regulation by cytokines IL-1beta, TNF-alpha, or IFN-gamma but released it when stimulated by CD40 ligand, a cytokine found in atheroma.

    CONCLUSIONS: These findings point to a novel role for the serine protease, neutrophil elastase, in matrix breakdown by macrophages, a critical process in adaptive remodeling of vessels and in the pathogenesis of arterial diseases.

  • 91.
    Domkin, Vladimir
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Chabes, Andrei
    Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS). Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Phosphines are ribonucleotide reductase reductants that act via C-terminal cysteines similar to thioredoxins and glutaredoxins2014In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 4, p. 5539-Article in journal (Refereed)
    Abstract [en]

    Ribonucleotide reductases (RNRs) catalyze the formation of 2'-deoxyribonucleotides. Each polypeptide of the large subunit of eukaryotic RNRs contains two redox-active cysteine pairs, one in the active site and the other at the C-terminus. In each catalytic cycle, the active-site disulfide is reduced by the C-terminal cysteine pair, which in turn is reduced by thioredoxins or glutaredoxins. Dithiols such as DTT are used in RNR studies instead of the thioredoxin or glutaredoxin systems. DTT can directly reduce the disulfide in the active site and does not require the C-terminal cysteines for RNR activity. Here we demonstrate that the phosphines tris(2-carboxyethyl)phosphine (TCEP) and tris(3-hydroxypropyl)phosphine (THP) are efficient non-thiol RNR reductants, but in contrast to the dithiols DTT, bis(2-mercaptoethyl)sulfone (BMS), and (S)-(1,4-dithiobutyl)-2-amine (DTBA) they act specifically via the C-terminal disulfide in a manner similar to thioredoxin and glutaredoxin. The simultaneous use of phosphines and dithiols results in ~3-fold higher activity compared to what is achieved when either type of reductant is used alone. This surprising effect can be explained by the concerted action of dithiols on the active-site cysteines and phosphines on the C-terminal cysteines. As non-thiol and non-protein reductants, phosphines can be used to differentiate between the redox-active cysteine pairs in RNRs.

  • 92.
    Drechsler, Michal
    Karlstad University, Faculty of Technology and Science.
    Models in chemistry education: A study of teaching and learning acids and bases in Swedish upper secondary schools2007Doctoral thesis, comprehensive summary (Other scientific)
    Abstract [en]

    This thesis reports an investigation of how acid-base models are taught and understood in Swedish upper secondary school. The definition of the concepts of acids and bases has evolved from a phenomenological level to an abstract (particle) level. Several models of acids and bases are introduced in Swedish secondary school. Among them an ancient model, the Arrhenius model and the Brønsted model. The aim of this study was to determine how teachers handle these models in their teaching. Further, to investigate Swedish upper secondary students’ ideas about the role of chemistry models, in general, and more specific, of models of acids and bases. The study consisted of two parts. First, a study was performed to get an overview of how acids and bases are taught and understood in Swedish upper secondary schools. It consisted of three steps: (i) the most widely used chemistry textbooks for upper secondary school in Sweden were analysed, (ii) six chemistry teachers were interviewed, and, (iii) finally also seven upper secondary school students were interviewed. The results from this study were used in the second part which consisted of two steps: (i) nine chemistry teachers were interviewed regarding their pedagogical content knowledge (PCK) of teaching acids and bases, and (ii) a questionnaire was administered among chemistry teachers of 441 upper secondary schools in Sweden. The results from the interviews show that only a few teachers chose to emphasise the different models of acids and bases. Most of the teachers thought it was sufficient to distinguish clearly between the phenomenological level and the particle level. In the analysis of the questionnaire three subgroups of teachers were identified. Swedish upper secondary chemistry teachers, on the whole, had a strong belief in the Brønsted model of acids and bases. However, in subgroup one (47 %) teachers’ knowledge of how the Brønsted model differs from older models was limited and diverse. Teachers in subgroup two (38 %) and three (15 %) seemed to understand the differences between the Brønsted model and older models, but teachers in subgroup 2 did not explain the history of the development of acids and bases in their teaching. Instead they (as teachers in subgroup one) relied more on the content in the textbooks than teachers in the third subgroup. Implications for textbook writers, teaching, and further research are discussed.

  • 93.
    Dubois, Louise
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Andersson, Karl
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Asplund, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Björkelund, Hanna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Radiology, Oncology and Radiation Science, Biomedical Radiation Sciences.
    Evaluating real-time immunohistochemistry on multiple tissue samples, multiple targets and multiple antibody labeling methods2013In: BMC Research Notes, ISSN 1756-0500, E-ISSN 1756-0500, Vol. 6, p. 542-Article in journal (Refereed)
    Abstract [en]

    Background

    Immunohistochemistry (IHC) is a well-established method for the analysis of protein expression in tissue specimens and constitutes one of the most common methods performed in pathology laboratories worldwide. However, IHC is a multi-layered method based on subjective estimations and differences in staining and interpretation has been observed between facilities, suggesting that the analysis of proteins on tissue would benefit from protocol optimization and standardization. Here we describe how the emerging and operator independent tool of real-time immunohistochemistry (RT-IHC) reveals a time resolved description of antibody interacting with target protein in formalin fixed paraffin embedded tissue. The aim was to understand the technical aspects of RT-IHC, regarding generalization of the concept and to what extent it can be considered a quantitative method.

    Results

    Three different antibodies labeled with fluorescent or radioactive labels were applied on nine different tissue samples from either human or mouse, and the results for all RT-IHC analyses distinctly show that the method is generally applicable. The collected binding curves showed that the majority of the antibody-antigen interactions did not reach equilibrium within 3 hours, suggesting that standardized protocols for immunohistochemistry are sometimes inadequately optimized. The impact of tissue size and thickness as well as the position of the section on the glass petri dish was assessed in order for practical details to be further elucidated for this emerging technique. Size and location was found to affect signal magnitude to a larger extent than thickness, but the signal from all measurements were still sufficient to trace the curvature. The curvature, representing the kinetics of the interaction, was independent of thickness, size and position and may be a promising parameter for the evaluation of e.g. biopsy sections of different sizes.

    Conclusions

    It was found that RT-IHC can be used for the evaluation of a number of different antibodies and tissue types, rendering it a general method. We believe that by following interactions over time during the development of conventional IHC assays, it becomes possible to better understand the different processes applied in conventional IHC, leading to optimized assay protocols with improved sensitivity.

  • 94.
    Dziedzic, Pawel
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Amino acid-catalyzed synthesis of amino acid derivatives: Application and semi-synthesis of Paclitaxel, Docetaxel and their derivatives2010Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    This thesis deals with different applications of organocatalysis, where amino acid derivatives and small peptides are applied as catalysts. First, the development of environmentally friendly aldol reactions, carried out in aqueous media is illustrated. The corresponding β-hydroxy ketones are formed with ee´s up to 99%. Chapter 3 describes the ability of β3-amino acids to selectively catalyze Mannich-type reactions and govern the formation of products with high anti-selectivity (up to >19:1) and ee´s up to 99%. In the following chapter, an amino acid-catalyzed one-pot three component Mannich reaction between dihydroxyacetone and PMP-protected imines, is presented. The corresponding a,a’-dihydroxy-b-aminoketones are obtained in high yields and with 82-95% ee. Next, an aza-Morita-Baylis-Hillman reaction was investigated where L-proline is the catalyst. The reaction proceeds with excellent chemo- and enantioselectivity to give the corresponding compounds in good yields and with 97-99% ee. Finally, the last part describes development of a proline-catalyzed Mannich reation between N-acyl imines and protected α-hydroxyaldehyes, providing access to different α-hydroxy-β-amino acids in good yields and high enantioselctivity (92-99% ee). The obtained amino acids were further applied in the semisynthesis of paclitaxel and docetaxel derivatives.

  • 95.
    Dziedzic, Pawel
    et al.
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Weibiao, Zou
    Hafrén, Jonas
    Córdova, Armando
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    The small peptide-catalyzed direct asymmetric aldol reaction in water2006In: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, Vol. 4, p. 38-40Article in journal (Refereed)
  • 96.
    Dziedzic, Pawel
    et al.
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Zhao, Gui-ling
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Córdova, Armando
    Stockholm University, Faculty of Science, Department of Organic Chemistry.
    Practical amino acid-catalyzed asymmetric synthesis pf protected α-hydroxy-amino aldehydes and acidsManuscript (preprint) (Other academic)
  • 97.
    Edgren, Ellen
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Analys av vitamin B12 i tillagad och återuppvärmd lax genom bioassay med Lactobacillus delbrueckii subart lactis ATCC® 7830TM2010Independent thesis Advanced level (degree of Master (One Year)), 20 credits / 30 HE creditsStudent thesis
    Abstract [sv]

    Halten näringsämnen i livsmedel varierar bland annat med tillagningsmetod, lagringstid och lagringsförhållanden, som exponering för ljus och syre. Oklarheter finns för mikrovågors inverkan på näringsämnen och framförallt på vitamin B12. Eftersom många äldre drabbas av brist på vitamin B12, är detta av intresse eftersom många hemmaboende äldre människor får hemleverans av färdiga matportioner som är avsedda att värmas, ofta i mikrovågsugn. Syftet med studien var att kunna ge en indikation på om uppvärmning genom mikrovågor påverkar halten vitamin B12 i färdiga kylda måltider. Detta gjordes genom litteratursammanställning om vitamin B12 och mikrovågar samt genom bioassay med Lactobacillus delbrueckii. Vitamin B12 i ouppvärmd lax, lax värmd i mikrovågsugn samt lax värmd i konventionell ugn analyserades. Tillväxten av bakterier uppskattades genom mätning av turbiditet. Högst vitamin B12-halt sågs i mikrovågsvärmd lax, därefter ouppvärmd lax och ugnsvärmd lax. Statistiskt signifikanta skillnader sågs för analyserade vitamin B12-halter mellan ouppvärmt- och ugnsvärmt prov respektive mellan mikrovågsvärmt- och ugnsvärmt prov. De varierande vitaminhalterna kan bero på metodfel, otillräcklig vitaminextraktion eller att bakterierna inte konsumerat allt tillgängligt vitamin vid turbiditetsmätning. Slutsatsen av studien är att halten vitamin B12 i livsmedel inte verkar påverkas negativt av mikrovågor, däremot finns tendenser att halten minskar beroende av temperatur och tillagningstid.

  • 98.
    Edlund, Bror
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Andersson, Jill
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Titanji, Vincent
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Dahlqvist, Ulla
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Ek, Pia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Zetterqvist, Örjan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Engström, Lorentz
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical and Physiological Chemistry.
    Amino acid sequence at the phosphorylated site of rat liver pyruvate kinase1975In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 67, no 4, p. 1516-1521Article in journal (Refereed)
    Abstract [en]

    One dominating peptic phosphopeptide, Asx-Thr-Lys-Gly-Pro-Glx-Ile-Glx-Thr-Gly-Val-Leu-Arg-Arg-Ala-(32P)SerP-Val-Ala-Glx-Leu, was obtained from rat liver pyruvate kinase (type L) phosphorylated by cyclic 3′,5′-AMP-stimulated protein kinase from the same tissue. The sequence around the phosphorylated serine residue is similar to that of a corresponding but smaller peptic phosphopeptide previously isolated from pig liver (type L) pyruvate kinase, Leu-Arg-Arg-Ala-(32P)SerP-Leu.

  • 99.
    Edlund, Karolina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular and Morphological Pathology.
    Molecular Characterisation and Prognostic Biomarker Discovery in Human Non-Small Cell Lung Cancer2012Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Non-small cell lung cancer (NSCLC) constitutes a clinically, histologically, and genetically heterogeneous disease entity that represents a major cause of cancer-related death. Early-stage patients, who undergo surgery with curative intent, experience high recurrence rates and the effect of adjuvant treatment is modest. Prognostic biomarkers would be of particular relevance to guide intensified treatment depending on expected outcome and moreover often infer a biological role in tumourigenesis.

    This thesis presents a translational study approach to establish a well-characterised NSCLC frozen-tissue cohort and to obtain a profile of each specimen with regard to genome-wide copy number alterations, global gene expression levels and somatic mutations in selected cancer-related genes. Furthermore, the generation of a formalin-fixed, paraffin-embedded tissue microarray enabled validation of findings on the protein level using immunohistochemistry. The comprehensive molecular characterisation, combined with data on clinical parameters, enabled the analysis of biomarkers linked to disease outcome. In Paper I, single nucleotide polymorphism arrays were applied to assess copy number alterations in NSCLC and associations with overall survival in adenocarcinoma and squamous cell carcinoma were described. In Paper II, we evaluated expression levels of selected stromal proteins in NSCLC using immunohistochemistry and the adhesion molecule CD99 was identified as an outcome-related biomarker in two independent cohorts. Paper III presents a strategy for prognostic biomarker discovery based on gene expression profiling, meta-analysis, and validation of protein expression on tissue microarrays, and suggests the putative tumour suppressor CADM1 as a candidate biomarker. In Paper IV, we propose a prognostic role for tumour-infiltrating IGKC-expressing plasma cells in the local tumour microenvironment, indicating an involvement of the humoral immune response in anti-tumor activity. In Paper V, we combined next-generation deep sequencing with statistical analysis of the TP53 database to define novel parameters for database curation.

    In summary, this thesis exemplifies the benefits of a translational study approach, based on a comprehensive tumour characterisation, and describes molecular markers associated with clinical outcome in NSCLC.

  • 100.
    Ek, Pia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Dahlqvist, Ulla
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Humble, Elisabet
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Engström, Lorentz
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Comparative kinetic studies on the L-type pyruvate kinase from rat liver and the enzyme phosphorylated by cyclic 3´, 5´-AMP-stimulated protein kinase1976In: Biochimica et Biophysica Acta, ISSN 0006-3002, E-ISSN 1878-2434, Vol. 429, no 2, p. 374-382Article in journal (Refereed)
    Abstract [en]

    The kinetics of rat liver L-type pyruvate kinase (EC 2.7.1.40), phosphorylated with cyclic AMP-stimulated protein kinase from the same source, and the unphosphorylated enzyme have been compared. The effects of pH and various concentrations of substrates, Mg2+, K+ and modifiers were studied. In the absence of fructose 1, 6-diphosphate at pH 7.3, the phosphorylated pyruvate kinase appeared to have a lower affinity for phosphoenolpyruvate (K0.5=0.8 mM) than the unphosphorylated enzyme (K0.5=0.3 mM). The enzyme activity vs. phosphoenolpyruvate concentration curve was more sigmoidal for the phosphorylated enzyme with a Hill coefficient of 2.6 compared to 1.6 for the unphosphorylated enzyme. Fructose 1, 6-diphosphate increased the apparent affinity of both enzyme forms for phosphoenolpyruvate. At saturating concentrations of this activator, the kinetics of both enzyme forms were transformed to approximately the same hyperbolic curve, with a Hill coefficient of 1.0 and K0.5 of about 0.04 mM for phosphoenolpyruvate. The apparent affinity of the enzyme for fructose 1, 6-diphosphate was high at 0.2 mM phosphoenolpyruvate with a K0.5=0.06 muM for the unphosphorylated pyruvate kinase and 0.13 muM for the phosphorylated enzyme. However, in the presence of 0.5 mM alanine plus 1.5 mM ATP, a higher fructose 1, 6-diphosphate concentration was needed for activation, with K0.5 of 0.4 muM for the unphosphorylated enzyme and of 1.4 muM for the phosphorylated enzyme. The results obtained strongly indicate that phosphorylation of pyruvate kinase may also inhibit the enzyme in vivo. Such an inhibition should be important during gluconeogenesis.

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