Bioelectrocatalytic systems are based on biological entities, such as enzymes, whole cells, parts of cells or tissues, which catalyse electrochemical processes that involve the interaction between chemical change and electrical energy. In all cases, biocatalysis is implemented by enzymes, isolated or residing inside cells or part of cells. Electron transfer (ET) phenomena, within the protein molecules and between biological redox systems and electronics, enable the development of various bioelectrocatalytic systems, which can be used both for fundamental investigations of enzymatic biological processes by electrochemical methods and for applied purposes, such as power generation, bioremediation, chemical synthesis and biosensing. Electrical communication between the biocatalyst’s redox centre and an electrode is essential for the functioning of the system. This can be established using two main mechanisms: indirect ET and direct ET. The efficiency of the ET influences important parameters such as the turnover rate of the biocatalyst, the generated current density and partially the stability of the system, which in their turn determine response time, sensitivity, detection limit and operational stability of biosensing devices or the power densities and current output of biofuel cells, and hence should be carefully considered when designing bioelectrocatalytic systems. This thesis focuses on approaches that facilitate ET in bioelectrocatalytic systems based on indirect and direct ET mechanisms. Both fundamental aspects of ET in bioelectrocatalytic systems and applications of such systems for biosensing and power generation are considered. First, a new hydrophobic mediator for oxidases – unsubstituted phenothiazine and its improved ET properties in comparison with commonly used mediators are discussed. Application of the mediator in electrochemical biosensors is demonstrated by glucose, lactate and cholesterol sensing. Utilisation of mediated biocatalytic cholesterol oxidation, as the anodic reaction for the construction of a biofuel cell acting as a power supply and an analytical device at the same time, is investigated to deliver a selfpowered biosensor. Also the enhancement of mediated bioelectrocatalysis by employment of microelectrodes as a transducer is examined. The effect of surface roughness on the current response of the microelectrodes under conditions of convergent diffusion is considered. The applicability of the laccase-based system for total phenol analysis of weakly supported water is demonstrated. Finally, a new electrochemical approach derived from collision-based electrochemistry applicable for examination of the ET process of a single enzyme molecule is described. All together, the results presented in this thesis contribute to the solution of the ‘electronic coupling problem’, arising when interfacing biomolecules with electronics and limiting the performance of bioelectrocatalytic systems in specific applications. The developed methods to facilitate ET will hopefully promote future biosensing devices and biofuel cells. I believe the new approach for investigation of ET processes at a single enzyme molecule will complement existing single molecule techniques, giving further insights into enzymatic ET mechanisms at the molecular level and filling the gap between fundamental understanding of biocatalytic processes and their potential for bioenergy production.

This thesis is an investigation into the hydrogenous behavior of Zintl phases. Zintl phases are comprised of an active metal (i.e alkali, alkaline earth, and rare earth) and a p-block element. The discussion gives an overview of the influence hydrogen affects the electronic and geometric structure of Zintl phases and subsequent properties. Incorporation of hydrogen into a Zintl phase is categorized as either polyanionic or interstitial Zintl phase hydrides. In the former the hydrogen covalently bonds to the polyanion and in the latter the hydrogen behaves hydridic, coordinates exclusively with the active metal, leading to an oxidation of the polyanion. Synthesis of hydrogenous Zintl phases may be through either a direct hydrogenation of a Zintl phase precursor or by combining active metal hydrides and p-block elements. The latter strategy typically leads to thermodynamically stable hydrides, whereas the former supports the formation of kinetically controlled products.  Polyanionic hydrides are exemplified by SrAlGeH and BaAlGeH. The underlying Zintl phases SrAlGe and BaAlGe have a structure that relates to the AlB2 structure type. These Zintl phases possess 9 valence electrons for bonding and, thus, are charge imbalanced species. Connected to the charge imbalance are superconductive properties (the Tc of SrAlGe and BaAlGe is 6.7 and 6.3 °C, respectively). In the polyanionic hydrides the hydrogen is covalently bonded as a terminating ligand to the Al atoms. The Al and Ge atoms in the anionic substructure [AlGeH]2- form corrugated hexagon layers. Thus, with respect to the underlying Zintl phases there is only a minimal change to the arrangement of metal atoms. However, the electronic properties are drastically changed since the Zintl phase hydrides are semiconductors.  Interstitial hydrides are exemplified by Ba3Si4Hx (1 < x < 2) which was obtained from the hydrogenation of the Zintl phase Ba3Si4. Ba3Si4 contains a Si46- “butterfly” polyanion. Hydrogenation resulted in a disordered hydride in which blocks of two competing tetragonal structures are intergrown. In the first structure the hydrogen is located inside Ba6 octahedra (I-Ba3Si4H), and in the second structure the hydrogen is located inside Ba5 square pyramids (P-Ba3Si4H2). In both scenarios the “butterfly anions appear oxidized and form Si44- tetrahedra. Hydrogenation may also be used as a synthesis technique to produce p-block element rich Zintl phases, such as silicide clathrates. During hydrogenation active metal is removed from the Zintl phase precursor as metal hydride. This process, called oxidative decomposition, was demonstrated with RbSi, KSi and NaSi. Hydrogenation yielded clathrate I at 300 °C and 500 °C for RbSi and KSi, respectively. Whereas a mixture of both clathrate I and II resulted at 500 °C for NaSi.  Low temperature hydrogenations of KSi and RbSi resulted in the formation of the silanides KSiH3 and RbSiH3. These silanides do not represent Zintl phase hydrides but are complex hydrides with discrete SiH3- complex species. KSiH3 and RbSiH3 occur dimorphic, with a disordered α-phase (room temperature; SG Fm-3m) and an ordered β-phase (below -70 °C; SG = Pnma (KSiH3); SG = P21/m ( RbSiH3)). During this thesis the vibrational properties of the silyl anion was characterized. The Si–H stretching force constants for the disordered α-phases are around 2.035 Ncm-1 whereas in the ordered b-forms this value is reduced to ~1.956 Ncm-1. The fact that SiH3- possesses stronger Si-H bonds in the α-phases was attributed to dynamic disorder where SiH3- moieties quasi freely rotate in a very weakly coordinating alkali metal ion environment.

A sustainable solution for meeting the energy demands at our planet is by utilizing wind-, solar-, wave-, thermal-, biomass- and hydroelectric power. These renewable and CO2 emission-free energy sources are highly variable in terms of spatial and temporal availability over the Earth, introducing the need for an appropriate method of storing and carrying energy. Hydrogen has gained significant attention as an energy storage- and carrier media because of the high energy density that is exploited within the ‘power-to-gas’ process chain. A robust way of producing sustainable hydrogen is via electrochemical water splitting. In this work the search for new heterogeneous catalyst materials with the aim of increasing energy efficiency in water splitting has involved methods of both electrochemical water splitting and chemical water oxidation. Some 21 compounds including metal- oxides, oxofluorides, oxochlorides, hydroxide and metals have been evaluated as catalysts. Two of these were synthesized directly onto conductive backbones by hydrothermal methods. Dedicated electrochemical cells were constructed for appropriate analysis of reactions, with one cell simulating an upscale unit accounting for realistic large scale applications; in this cell gaseous products are quantified by use of mass spectrometry. Parameters such as real time faradaic efficiency, production of H2 and O2 in relation to power input or overpotentials, Tafel slopes, exchange current density and electrochemical active surface area as well as turnover numbers and turnover frequencies have been evaluated. Solubility, possible side reactions, the role of the oxidation state of catalytically active elements and the nature of the outermost active surface layer of the catalyst are discussed. It was concluded that metal oxides are less efficient than metal based catalysts, both in terms of energy efficiency and in terms of electrode preparation methods intended for long time operation. The most efficient material was Ni-Fe hydroxide electrodeposited onto Ni metal foam as conductive backbone. Among the other catalysts, Co3Sb4O6F6 was of particular interest because the compound incorporate a metalloid (Sb) and redox inert F and yet show pronounced catalytic performance. In addition, performance of materials in water splitting catalysis has been discussed on the basis of results from electron microscopy, solubility experiments and X-ray diffraction data.

The use of enzymes as catalysts in organic synthesis constitutes an attractive alternative to conventional chemical catalysis. Enzymes are non-toxic and biodegradable and they can operate under mild reaction conditions. Furthermore, they often display high chemo-, regio- and stereoselectivity, enabling specific reactions with single product outcome. By the use of protein engineering, enzymes can be altered for the specific needs of the researcher. The major part of this thesis describes engineering of lipase A from Candida antarctica (CalA), for improved enantioselectivity in organic synthetic transformations. The first part of the thesis describes a highly combinatorial method for the introduction of mutation sites in an enzyme library. By the simultaneous introduction of nine mutations, we found an enzyme variant with five out of the nine possible mutations. This quintuple variant had an enlarged active site pocket and was enantioselective and active for our model substrate, an ibuprofen ester. This is a bulky substrate for which the wild-type enzyme shows no enantioselectivity and very poor activity. In the second part of the thesis, we continued our approach of combinatorial, focused enzyme libraries. This time we aimed at decreasing the alcohol pocket of CalA, in order to increase the enantioselectivity for small and medium-sized secondary alcohols. The enzyme library was bound on microtiter plates and screened by a transacylation reaction in organic solvent. This library yielded an enzyme variant with high enantioselectivity for the model substrate 1-phenyl ethanol, and high to excellent selectivity for other alcohols tested. Screening in organic solvent is advantageous since a potential hit is more synthetically useful. In the third part of the thesis, we used manipulated beads of controlled porosity glass (EziG™) for enzyme immobilization, and demonstrated the generality of this carrier for several enzyme classes. EziG™ allowed fast enzyme immobilization with simultaneous purification and yielded active biocatalysts in all cases. The last project describes the function of the proposed active site flap in CalA. In our study, we removed this motif. The engineered variant was compared to the wild-type enzyme by testing the amount of interfacial activation and the selectivity for certain alcohols. We showed that the motif is indeed controlling the entrance to the active site and that the flap is not part of the enantioselectivity determining machinery. / <p>At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.</p>

This thesis focuses on the development of methods for the activation of the hydroxyl group in non-derivatized alcohols in substitution reactions. The thesis is divided into two parts, describing three different catalytic systems. The first part of the thesis (Chapter 2) describes nucleophilic allylation of amines with allylic alcohols, using a palladium catalyst to generate unsymmetrical diallylated amines. The corresponding amines were further transformed by a one-pot ring-closing metathesis and aromatization reaction to afford β-substituted pyrroles with linear and branched alkyl, benzyl, and aryl groups in overall moderate to good yields. The second part (Chapters 3 and 4) describes the direct intramolecular stereospecific nucleophilic substitution of the hydroxyl group in enantioenriched alcohols by Lewis acid and Brønsted acid/base catalysis. In Chapter 3, the direct intramolecular substitution of non-derivatized alcohols has been developed using Fe(OTf)3 as catalyst. The hydroxyl groups of aryl, allyl, and alkyl alcohols were substituted by the attack of O- and N-centered nucleophiles, to provide five- and six-membered heterocycles in up to excellent yields with high enantiospecificities. Experimental studies showed that the reaction follows first-order dependence with respect to the catalyst, the internal nucleophile, and the internal electrophile of the substrate. Competition and catalyst-substrate interaction experiments demonstrated that this transformation proceeds via an SN2-type reaction pathway. In Chapter 4, a Brønsted acid/base catalyzed intramolecular substitution of non-derivatized alcohols was developed. The direct intramolecular and stereospecific substitution of different alcohols was successfully catalyzed by phosphinic acid (H3PO2). The hydroxyl groups of aryl, allyl, propargyl, and alkyl alcohols were substituted by O-, N-, and S-centered nucleophiles to generate five- and six-membered heterocycles in good to excellent yields with high enantiospecificities. Mechanistic studies (both experiments and density functional theory calculations) have been performed on the reaction forming five-membered heterocyclic compounds. Experimental studies showed that phosphinic acid does not promote SN1 reactivity. Rate-order determination indicated that the reaction follows first-order dependence with respect to the catalyst, the internal nucleophile, and the internal electrophile. DFT calculations corroborated with a reaction pathway in which the phosphinic acid has a dual activation mode and operates as a bifunctional Brønsted acid/Brønsted base to simultaneously activate both the nucleophile and nucleofuge, resulting in a unique bridging transition state in an SN2-type reaction mechanism. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: Manuscript. Paper 4: Manuscript.</p>

Perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) are two commonly found contaminants associated with various manufacturing products, such as firefighting foam, non-stick coatings, electronics and water repellants. These compounds are persistent, bioaccumulative, and toxic and may therefore pose a serious health risk to living organisms. Earlier studies have shown that PFOS and PFOA affected the fatty acid β-oxidation, i.e. the energy metabolism in liver. This study evaluates the effects of PFOS and PFOA on fatty acid metabolism in domestic chicken liver cells.  Liver tissues were obtained from chicken embryos treated in ovo with PFOS or with PFOA at low (0.1 µg/g) and high (1.0 and 1.6 µg/g) concentration levels. The fatty acids were extracted and derivatized into fatty acid methyl esters (FAMEs). The analysis was conducted by gas chromatography coupled with mass spectrometry. Results showed that a lower concentration of PFOS and a lower percentage of DMSO significantly affected the concentrations of fatty acids in livers of chicken embryos. PFOA-treated samples also showed some significant elevated fatty acid concentrations. Almost all fatty acid concentrations of treated liver samples exceeded the concentrations of the untreated samples.

Electron crystallography is complementary to X-ray crystallography. Single crystal X-ray diffraction requires the size of a crystal to be larger than about 5 × 5 × 5 μm3 while a TEM allows a million times smaller crystals being studied. This advantage of electron crystallography has been used to solve new structures of small crystals. One method which has been used to collect electron diffraction data is rotation electron diffraction (RED) developed at Stockholm University. The RED method combines the goniometer tilt and beam tilt in a TEM to achieve 3D electron diffraction data. Using a high angle tilt sample holder, RED data can be collected to cover a tilt range of up to 140o.  Here the crystal structures of several different compounds have been determined using RED. The structure of needle-like crystals on the surface of NiMH particles was solved as La(OH)2. A structure model of metal-organic layers has been built based on RED data. A 3D MOF structure was solved from RED data. Two halide perovskite structures and two newly synthesized aluminophosphate structures were solved. For those beam sensitive crystals characterized here, sample cooling down to -170oC was used to reduce the beam damage. The low temperature not only reduces electron beam damage, but also keeps the structure more stable in the high vacuum in a TEM and improves the quality of the diffraction data. It is shown that cooling can improve the resolution of diffraction data for MOFs and zeolites, for samples undergoing phase changes at low temperature, the data quality could be worse by cooling. In summary, cooling can improve the ED data quality as long as the low temperature does not trigger structural changes. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 1: Manuscript. Paper 3: Submitted.</p>

The aim of this study was to examine if perfluorinated alkylated substances (PFASs) reaches north of Sweden by long-range atmospheric transport. This was done by monitoring the levels of PFASs in reindeer livers at two locations in 2002 and 2010, respectively. The reindeers have lived all of their lives in the mountains and therefore the main source of exposure for PFASs is through air. The samples were extracted and analysed for 24 different PFASs using ultra performance liquid chromatography tandem mass spectrometer (UPLC-MS/MS). The most significant change concerns perfluorooctane sulfonic acid (PFOS) which decreased significantly from 6.1 ng/g at the most northern location (Ammarnäs/Biergenis) in 2002 to 0.87 ng/g 2010. At the other sampling location, Glen, PFOS decreased from 5.0 to 3.2 ng/g during the eight years. Mainly PFOS and longer chain carboxylates were found. The results revealed that the levels of many compounds decreased in time. The location seems to have an impact on the level of perfluorinated compounds present and most likely the distribution of them in the air, since certain PFASs have increased and decreased differently in time between the two locations. Since PFASs are non-volatile, they are believed to be degradation products of volatile compounds such as fluorotelomer alcohols (FTOHs) and perfluoroalkylated sulfonamido alcohols (FOSEs). FTOHs and FOSEs are released, translocated by long-range atmospheric transport and degraded to perfluorinated compounds in organisms or atmosphere.

Persistent organic pollutants such as polychlorinated biphenyls (PCBs), DDT and polybrominated diphenyl ethers (PBDEs) tend to accumulate in biota and are transferred through the aquatic food web, which result in a high accumulation in marine mammals. In recent years various novel flame retardants (nBFRs), which have replaced the banned PBDEs, have also started to occur in the environment. These nBFRs have similar properties as PBDEs, such as long-range transport and accumulation in biota. The purpose with this study was to evaluate a method by using pre-packed silica columns for quantification of PCBs, DDT, PBDEs and nBFRs in seal blubber, in order to facilitate the pre-treatment and decrease the time. To elute the different POPs from the pre-packed silica column; hexane, toluene and dichloromethane were used in different stages. By using this method levels of PCB and DDT were determined. For DDT the concentration was 8.28 ng/g lipid and 8.94 ng/g lipid for the two samples that was analysed, and the analysis of the PCBs showed a higher trend for the higher chlorinated PCBs. As the pre-packed silica columns are a relative new method. Further studies are therefore needed on these columns to further improve the sample clean-up and fractionation of the different POPs in environmental samples.

Per-and polyfluoroalkyl substances (PFAS) are anthropogenic compounds that have been spread all over the world. The use of fluorotelomer compounds, short-chained homologues, and other PFASs with perfluorinated moieties has emerged recent years. One of these emerging compound classes is polyfluoroalkyl phosphate esters (PAPs), which have the ability to degrade into persistent PFCAs. The aim of this thesis was to assess the contribution of PAPs and other precursors to the exposure of PFCAs to humans and the environment. The main objective was to analyze a wide range of PFAS in human serum, wild bird eggs, indoor dust, waste water, and sludge. There was a significant contribution from selected precursors to the total amount of PFASs in the abiotic compartments indoor dust, waste water, and sludge. Levels of PAPs found in house dust exceeded those of PFCAs and perfluorosulfonic acids (PFSAs), revealing PAPs as a world-wide important exposure source. A net increase was during waste water treatment was observed for several PFASs in Swedish waste water treatment plants. Together with presence of precursor compounds and intermediates in the influent water and the sludge, this suggest that degradation of PFCA precursors contributed to the increase of PFCAs. Detection of precursors in human serum, together with slow declining trends of PFCAs, revealed an ongoing exposure of PFCAs to the general population of Australia. The diPAPs and the FTSAs were also detected in raptor bird eggs from Sweden from both the terrestrial and the freshwater environment. The precursors concentrations and patterns observed reveal that current regulatory measures are insufficient for the purpose of protecting humans and the environment from PFASs exposure.