A new method for analysis of acidic herbicides, mostly phenoxy acids and their esters, in cereals with liquid chromatography-tandem quadrupole mass spectrometry (LS-MS/MS) has been developed. Samples were hydrolyzed with sodium hydroxide in order to release covalently bound compounds followed by neutralization and finally extraction with acidified ethyl acetate. The extraction efficiency for both ester formulations and acids were studied. Acceptable results (70-120 %) were obtained for 2,4-D, dichlorprop, MCPA and mecoprop for both esters and acids. However, low recoveries were observed for ester formulations of dicamba, fluroxypyr, fluazifop and haloxyfop, possibly due to the complex structure of the compounds in combination with the matrix and/or incomplete hydrolysis step. The limit of quantification (LOQ) for targeted pesticides was 0.01 mg/kg. The method has been tested in the EU Proficiency Test for cereals with good results.

There is convincing research which shows that the enzyme alkaline phosphatase (ALP) has a central role in the mineralization of bone, more precisely that its catalytic activity is needed in the process. ALP is found on the surface of matrix vesicles where the mineral is formed. One theory about the function of the enzyme is that it binds to fibrous collagen in the bone and thereby incorporating the mineral into the bone. The purpose of this study is to establish whether ALP binds to collagen. If this is the case, more elaborate studies around this will be performed. The strength of the binding between collagen and the different types of ALP will be evaluated, as well as on which part of the collagen the binding occurs. The binding is going to be studied by constructing a method for the ÄKTApurifier system.   Initially, the pureness of the different type of collagens was determined by using SDS-PAGE and the activity of the different types of ALP was established. These were also compared with a native PAGE. In SDS-PAGE, bovine type I collagen showed markings for a triple helix, a double helix and two single strains, α1 and α2. Bovine type II collagen showed markings for a double helix and α1-strains. Human type I collagen showed markings for a triple helix, two double helixes, two α-strains and contaminations. Trials with collagen in Native PAGE did not provide any results. However, the trials with ALP revealed that the different types of ALP had different charge.   Thereafter, blotting was performed. The results showed that all the different types of ALP, besides from E. coli, binds to bovine collagen type I and II and human collagen type I, however within various periods of time. In the trials with collagen coated plates the acquired results showed that some of the different types of ALP bind to collagen. ALP from liver binds the strongest to both collagen type I from rat and type IV from mouse. Intestinal ALP also binds to both types of collagen but not nearly as strong as liver ALP. Serum from rats did bind to collagen type I from rat but not to collagen type IV from mouse. ALP from kidney and human serum did not bind to either types of collagen. The trials concerning the ÄKTApurifier system were executed with ALP from liver alone because it had been proven to bind to bovine type I collagen through the previous methods. The results confirmed that ALP from liver binds to this type of collagen.   The conclusions from this study are that ALP does indeed bind to collagen and does so to the triple helix and double helix form as well as the single strains of collagen. In other words the part of the structure in collagen that ALP binds to must exist in all three stages of collagen formation. Furthermore, it seems like some of the different types of ALP has a higher affinity for binding to collagen, as the time for binding to collagen varies for the different types of ALP. The results differed between methods concerning different types of ALP. Although, the method we consider to give the best result was blotting. However, the method using ÄKTApurifier can be complementary but needs further development.

Today, a majority of the world’s energy need is supplied through sources that are finite and, at the current usage rates, will be consumed shortly. The high energy demand and pollution problems caused by the widespread use of fossil fuels make it increasingly necessary to develop renewable energy sources of limitless duration with smaller environmental impact than the traditional energy sources. Three fuels – rapeseed methyl ester (RME), Fischer-Tropsch (FT) diesel and FT jet fuel – derived from biomass, coal or gas were evaluated in this project. The fuel properties evaluated are in most cases listed in standards, often with recommendations, developed for biodiesel, petroleum diesel and jet fuel. Biodiesel is monoalkyl esters, e.g. RME, produced by transesterification of triglycerides in vegetable oil and an alcohol to esters and glycerin. This produce a fuel that is suitable as a direct substitution for petroleum diesel. Biodiesel may be used in pure form or in a blend with petrodiesel. Oxidative degradation and weak low temperature performance of biodiesel are properties of concern when substituting petrodiesel with biodiesel, as was shown in this project. The experiments show that oxidative stability can be improved with a synthetic antioxidant, e.g. butylated hydroxytoluene (BHT). The FT process converts syngas (a mixture of hydrogen and carbon monoxide) to a range of hydrocarbons. Syngas can be generated from a variety of carbon sources, e.g. coal, natural gas and biomass. The high-temperature (300-350 °C) FT process with iron-based catalysts is used for the production of gasoline and linear low molecular mass olefins (alkenes). The lowtemperature (200-240 °C) FT process with either iron or cobalt catalysts is used for the production of high molecular mass linear waxes. By applying various downstream processes, fuels suitable for substitution of petrodiesel and conventional jet fuel can be obtained. The FT fuels have lower densities than the conventional fuels. However, conclusions from this project are that most of the properties of FT fuels are better, or equal, than conventional petroleum fuels.

<p>Today, a majority of the world’s energy need is supplied through sources that are finite and, at the current usage rates, will be consumed shortly. The high energy demand and pollution problems caused by the widespread use of fossil fuels make it increasingly necessary to develop renewable energy sources of limitless duration with smaller environmental impact than the traditional energy sources.</p><p>Three fuels – rapeseed methyl ester (RME), Fischer-Tropsch (FT) diesel and FT jet fuel – derived from biomass, coal or gas were evaluated in this project. The fuel properties evaluated are in most cases listed in standards, often with recommendations, developed for biodiesel, petroleum diesel and jet fuel.</p><p>Biodiesel is monoalkyl esters, e.g. RME, produced by transesterification of triglycerides in vegetable oil and an alcohol to esters and glycerin. This produce a fuel that is suitable as a direct substitution for petroleum diesel. Biodiesel may be used in pure form or in a blend with petrodiesel. Oxidative degradation and weak low temperature performance of biodiesel are properties of concern when substituting petrodiesel with biodiesel, as was shown in this project. The experiments show that oxidative stability can be improved with a synthetic antioxidant, e.g. butylated hydroxytoluene (BHT).</p><p>The FT process converts syngas (a mixture of hydrogen and carbon monoxide) to a range of hydrocarbons. Syngas can be generated from a variety of carbon sources, e.g. coal, natural gas and biomass. The high-temperature (300-350 °C) FT process with iron-based catalysts is used for the production of gasoline and linear low molecular mass olefins (alkenes). The lowtemperature (200-240 °C) FT process with either iron or cobalt catalysts is used for the production of high molecular mass linear waxes. By applying various downstream processes, fuels suitable for substitution of petrodiesel and conventional jet fuel can be obtained. The FT fuels have lower densities than the conventional fuels. However, conclusions from this project are that most of the properties of FT fuels are better, or equal, than conventional petroleum fuels.</p>

Organophosphate and phthalate esters are polymer additives that are frequently found in air and dust in indoor environments. This thesis describes the development and application of air sampling and selective mass spectrometric (MS) determination of these two compound groups. It also describes the sampling and screening of these compounds in indoor air and dust from different environments and the development and evaluation of a method for the extraction and determination of these compounds in indoor dust. An air sampling method previously used for air sampling of only organophosphate esters, is here demonstrated to be applicable for simultaneous sampling of both phthalate and organophosphate esters. Selective detection using tandem mass spectrometry (MS/MS) showed good results for simultaneous determination of organophosphate and phthalate esters in air. The high selectivity of this technique was especially advantageous when analyzing dust. Comparison of chemical profiles of the organophosphate ester between indoor environments including daycare centers, offices and private homes differed between the types of locations while the phthalate profiles were rather similar. Comparison of concentration levels of these compounds in multi-storey apartment buildings classified as either high or low risk “sick” buildings could not differentiate the two classes of buildings. Further, the studies also points out some potential sources of organophosphate and phthalate esters in these indoor environments. In general the levels of phthalate esters were consistently higher than the levels of organophosphate esters both in air and in dust. Phthalate and organophosphate esters were also determined in a dust standard reference material; seventeen of the targeted compounds were quantified out of which ten had not previously been reported for this reference material. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 3: Submitted. Paper 4: Manuscript.</p>

Stärkelse används vid papperstillverkning för att öka styrkeegenskaperna hos pappret. Denna stärkelse kan av olika anledningar brytas ned och bli sämre, då främst på grund av bakterier. Detta kan leda till försämrad papperskvalitet och i värsta fall kassationer av slutprodukten. För att undersöka om stärkelsen blivit angripen av bakterier kan man göra en bakterieodling. Nackdelen med detta är att det tar väldigt lång tid innan man ser något resultat. En annan effekt av bakterierna är att stärkelsens viskositet blir lägre, dvs. vätskan blir mindre trögflytande. Detta är en parameter som kan mätas med en viskometer. Syftet är att utarbeta en ny mätstandard för att detektera bakteriella angrepp, där den främsta fördelen är att mätmetoden ger en snabb och säker respons och därmed minskar reaktionstiden för att motverka bakterierna. Målet med examensarbetet är att mäta viskositeten på stärkelse kokad i bruket samt på stärkelse kokad på labb och därmed börja bygga upp en databas med värden för att validera viskometern. Arbetet har utförts i laboratoriet på Tech Center vid Smurfit Kappa, Piteå. En viktig upptäckt som gjordes var att pH-värdet kan vara lågt men det betyder nödvändigtvis inte att man har ett bakterieangrepp. Arbetet begränsas väldigt mycket av tidsperioden på fem veckor och det är möjligt att fler mätningar skulle ge ett annat resultat. De mätningar som gjorts tyder dock på att det går att mäta viskositeten på stärkelsen dag till dag och att parametern kommer att vara oförändrad så länge koncentrationen är konstant. Arbetet med insamling av mätdata från labbkokad stärkelse samt stärkelse kokad i jetkokare på bruket är en början till att validera metoden. Fler mätningar kommer att utföras för att kunna föra statistik på viskositeten. Punkter som temperatur och koncentrationsgradienters effekt på viskositeten kommer att utvärderas. Målet är att kunna korrigera för dessa så att man i slutändan ska kunna korrelera data till bakteriella angrepp. Om detta lyckas är nästa steg att implementera metoden som en standard på laboratoriet. / Starch is used in papermaking to increase the strength properties of the paper. The starch can for various reasons degrade and become worse, primarily due to bacteria. This can lead to poor paper quality and in worst case to rejects of the final product. To investigate whether the starch has become infested with bacteria, a bacteria culture can be made. The downside of this is that it takes very long time before any results can be seen. Another effect of the bacteria is that the viscosity of the starch gets lower, i.e. the liquid becomes less viscous. This is a parameter that can be measured with a viscometer. The aim is to develop a new measurement standard for detecting bacterial attacks, where the main advantage is that the method provides a quick and reliable response, thus reducing the reaction time to counteract the bacteria. The goal of the project is to measure the viscosity of starch cooked in the mill and starch cooked in the lab and by doing so, start to build up a database of values to validate the viscometer. The work has been carried out in the laboratory at Tech Center at Smurfit Kappa, Piteå. An important discovery was made that the pH may be low but it does not necessarily have to be a bacterial attack. The work is limited a lot by the time period of five weeks and it is possible that more measurements would produce a different result. The measurements made suggest that it is possible to measure the viscosity of the starch from one day to another and that the parameter will remain unchanged as long as the concentration is constant. The work of collecting data from lab cooked starch and starch cooked in the jet boiler at the mill is the beginning to validate the method. More measurements will be carried out in order to keep statistics of the viscosity. Factors like temperature and concentration’s effects on the viscosity will be evaluated. The goal is to correct for these in order to be able to correlate data to a bacterial infestation. If successful, the next step is to implement the method as a standard in the laboratory.

In gas chromatography, analytes are separated by differences in their partition between a mobile phase and a stationary phase. Temperature-program, column dimensions, stationary and mobile phases, and flow rate are all parameters that can affect the quality of the separation in gas chromatography. To achieve a good separation (in a short amount of time) it is necessary to optimize these parameters. This can often be quite a tedious task. Using computer simulations, it is possible to both gain a better understanding of how the different parameters govern retention and separation of a given set of analytes, and to optimize the parameters within minutes. In the research presented here, this was achieved by taking a thermodynamic approach that used the two parameters ΔH (enthalpy change) and ΔS (entropy change) to predict retention times for gas chromatography. By determining these compound partition parameters, it was possible to predict retention times for analytes in temperature-programmed runs. This was achieved through the measurement of the retention times of n-alkanes, PAHs, alcohols, amines and compounds in the Grob calibration mixture in isothermal runs. The isothermally obtained partition coefficients, together with the column dimensions and specifications, were then used for computer simulation using in-house software. The two-parameter model was found to be both robust and precise and could be a useful tool for the prediction of retention times. It was shown that it is possible to calculate retention times with good precision and accuracy using this model. The relative differences between the predicted and experimental retention times for different compound groups were generally less than 1%. The scientific studies (Papers I-IV) are summarized and discussed in the main text of this thesis. / At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Submitted.

The human brain (lat. cerebrum) is the most complex and heterogeneous organ in the human body. It is involved in a great number of body functions like movement, touch sensing, vision, hearing, smelling, hormone regulation and many more. In no other organ, the molecular communication mechanisms between different cells are so poorly understood. Due to the extensive diversity of processes that are controlled by the brain, diseases and injuries of the nervous system affect the human body significantly. Because of the immense complexity of the brain, the molecular mechanisms underlying the pathology of the diseases remain largely unknown. Hence, there is an urgent need for the development of new analytical strategies in order to investigate these conditions on a molecular level. Here, a central focus lies in the study of protein and peptide expression profiles, which can provide an insight in ongoing molecular mechanisms underlying the pathophysiology of the diseases. A powerful approach for studying proteins and peptide dynamics is mass spectrometry based proteomics, which is defined as the comprehensive study of all proteins expressed in a biological matrix at a certain point of time. The central objective of this thesis was to develop and employ different mass spectrometric techniques to study protein and peptide dynamics in the central nervous system in different neurological diseases. The individual studies comprise different aspects of proteome research. The first two studies included clinical proteomic applications for investigating protein dynamics in traumatic brain injury and amyotrophic lateral sclerosis. A further study was focused on method development for MS analysis of intact neural cells. The final three projects described in this thesis comprised MS based protein and peptide imaging in brain and spinal cord tissue samples. Here, the aim was to elucidate topological changes in protein expression in ALS as well as neuropeptide alterations in distinct brain structures in L-DOPA induced dyskinesia (LID) in Parkinson’s disease. / Felaktigt tryckt som Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology 713

Liquid chromatography is an important separation technique for a vast number of analytes. This thesis mainly focuses on the development of stationary phases for liquid chromatography based on zwitterionic sulfobetaine polymers. In the thesis, various ways to prepare zwitterionic polymers in an aqueous environment using reversible addition fragmentation chain transfer (RAFT) polymerization are described. Both telomers, i.e. short soluble polymer chains containing a functional terminal group, as well as graft polymers on various supports have been synthesized. The RAFT polymerization technique provides an increased degree of control of the final polymers, which may aid in the preparation of more specifically tailored separation materials. Sulfobetaine polymers carry both a positive and a negative charge within a single entity, which results in interesting solution properties as well as highly biocompatible features. These unique features make them especially suited for separation of highly polar and/or charged compounds. An example of the successful separation of short peptides using a stationary phase synthesized with the RAFT technique is given. The unusual properties of sulfobetaine-type polymers are believed to be associated with the structure of water close to the polymer. A study of water structure in some silica based stationary phase grafted with zwitterionic sulfobetaine polymers was conducted. The impact of water structure on retention characteristics was investigated.

Gas chromatography (GC) has been utilized for the study of enantiomer resolution of the atropisomers of PCBs, o,p´-DDD and o,p´-DDT. Different substituents and concentrations of cyclodextrin, capillary dimensions and type of stationary phase films have been investigated to achieve the resolution of as many of the atropisomers on one column as possible. The results indicated that the butyl substitution of 6-hydroxyl and the methyl substitution of 2- and 3-hydroxyl were the most promising for the enantiomeric separation. Using Capillary Electrophoresis (CE), the trimers and monomers of PM-β-CDs were compared for enantiomeric resolution, as well as comparing the cationic PMMA-β-CD with the anionic HS-β-CD. In these studies the trimer did not show an improved resolution for mepivacaine, when compared to the equimolar concentration of the monomer. The cationic CD gave increased resolution values for ibuprofen when compared to the anionic CD. A scheme for reversing enantiomeric elution order of both the basic propranolol and acidic ibuprofen is also presented, with the aim of facilitating the detection of impurities in a high sample loading. The detection of 1% of each enantiomer of propranolol, and 1% of R(-)-ibuprofen, was demonstrated, with elution prior to the tailing peak of the corresponding enantiomer. Dimethylacrylamide-coated capillaries were used in this work, and the stability of this coating was demonstrated, giving a highly reduced electroosmotic flow for up to six months. Enantiomeric baseline separations of omeprazole and 5-hydroxyomeprazole have also been achieved with both CE and Open Tubular Capillary Electrochromatography (OT-CEC) methods. With CE-UV, both a non-aqueous method (using HDMS-β-CD) and an aqueous method (using HS-β-CD) were used for enantiomeric resolution of the two racemates. Resolution of omeprazole was also achieved using CE-Electrospray Ionization-Mass Spectrometry (ESI-MS). In OT-CEC, avidin was immobilized on the inside surface of a fused silica capillary and was employed as chiral selector for the enantiomeric baseline resolution of omeprazole and 5-hydroxyomeprazole.