| 1. |
- Aminlashgari, Nina, 1986-
(författare)
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SALDI-MS Method Development for Analysis of Pharmaceuticals and Polymer Degradation Products
- 2012
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Surface assisted laser desorption ionization-mass spectrometry (SALDI-MS) was evaluated as a new tool for analysis of polymer degradation products. A SALDI method was developed enabling rapid analysis of low molecular mass polyesters and their degradation products. In addition, the possibility to utilize nanocomposite films as easy-to-handle surfaces for analysis of pharmaceutical compounds was investigated. Poly(ε-caprolactone) was used as a model compound for SALDI-MS method development. The signal-to-noise values obtained by SALDI-MS were 20 times higher compared to traditional matrix assisted laser desorption ionization-mass spectrometry (MALDI-MS) of the same samples with 2,5-dihydroxybenzoic acid as a matrix. Halloysite nanoclay and magnesium oxide showed best potential as surfaces and clean backgrounds in the low mass range were observed. The SALDI-MS method for the analysis of polyester degradation products was also verified by electrospray ionization-mass spectrometry (ESI-MS). An advantage over ESI-MS is the possibility to directly analyze degradation products in buffer solutions. Compared to gas chromatography-mass spectrometry (GC-MS) it is possible to analyze polar compounds and larger molecular mass ranges at the same time as complicated extraction steps are avoided. The possibility to use nanocomposite films as surfaces instead of free nanoparticles was evaluated by solution casting of poly(lactide) (PLA) films with eight inorganic nanoparticles. The S/N values of the pharmaceutical compounds, acebutolol, propranolol and carbamazepine, analyzed on the nanocomposite surfaces were higher than the values obtained on the surface of plain PLA showing that the nanoparticles participated in the ionization/desorption process even when they are immobilized. Beside the ease of handling, the risk for instrument contamination is reduced when nanocomposites are used instead of free nanoparticles. The signal intensities depended on the type of drug, type and concentration of nanoparticle. PLA with 10 % titanium oxide or 10 % silicon nitride functioned best as SALDI-MS surfaces.
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| 2. |
- Karlsson, Maria
(författare)
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Protected Lignin Biorefining: Fundamental Insights on Lignin Reactivity
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The potential to utilize lignin, which constitutes as much as 15-30% of the biomass, needs further evaluation. In the transition towards a bioeconomy, lignin has the potential to replace fossil-based phenols. Attempts to valorize the available technical lignins are ongoing; however, that lignin suffers from molecular heterogeneity. Accordingly, new process concepts, coined in the term lignin biorefineries, are required to obtain less heterogenic lignin with different properties and preserved molecular structure.In this study, a new lignin extraction concept was investigated, where the structural properties of lignin were preserved to a high degree using a physical protection strategy. The principle of preserving the lignin structure was based on a cyclic organosolv extraction concept. At first, a two-step concept was evaluated, where a hydrothermal extraction was performed to recover hemicellulose, followed by a cyclic organosolv extraction to obtain the lignin. Trend studies were performed for the individual cycles to gain a deeper understanding of how the lignin structure was affected by the cycles. To further investigate the method, chemometrics and design of experiment were used to gain knowledge about how different properties of lignin were affected by the extraction conditions and how the properties of lignin could be tailored. Based on the knowledge from the chemometric study and the observations from the two-step method, a refined one-step method was developed to obtain lignin with further improved analytical quality, i.e., up to 95% of the interunit linkages could be assigned for spruce lignin by heteronuclearsingle quantum coherence (HSQC) nuclear magnetic resonance (NMR)spectroscopy, 13C NMR and size exclusion chromatography (SEC). The universality of the method was investigated for different wood species, such as spruce and birch. The results indicate the applicability of the concept using different raw materials.The complex nature of lignin substrates conveys the need for robust analytic techniques. Herein, NMR studies were complemented by matrix-assisted laser desorption/ionization (MALDI) time of flight (TOF) mass spectrometry (MS) to provide new insights into molecular lignin populations with respect to reactivity during the organosolv extraction.Finally, a proof of concept for an application was investigated. The cyclic organosolv extracted lignin was used in a fundamental study on lignin nanoparticles (LNP), together with benchmark technical lignins, to gain knowledge about the role of the molecular structure in the LNP properties. It is suggested that the molecular structure of lignin plays an important role in determining the size and morphology of LNPs, opening possibilities to molecularly tailor LNP properties.
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| 3. |
- Kwan, Isabella, 1994-
(författare)
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Bark Biorefinery : Isolation, Characterization and Application
- 2023
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- To fulfill the United Nations’ 17 sustainable development goals, there is a need to transition from a petroleum-based society toward a more sustainable one which requires new solutions and the production of materials, chemicals, and energy from renewable resources. Using side-stream products from industries to produce value-added products will be economically beneficial. Furthermore, finding more environmentally friendly process routes will aid industries in achieving their goal of reducing carbon dioxide emissions and contributing to a more sustainable society.Tree bark is the outer protective layer of a tree and today, mills and factories incinerate the bark to produce energy. Norway spruce (Picea abies) bark consistsof cellulose, lignin, hemicellulose, and extractives. Applying the biorefinery concept makes it possible to extract and isolate these compounds in the bark and utilize them to produce high-value materials and chemicals.This thesis applies the biorefinery concept to isolate cellulose and lignin from spruce bark by using mild extraction processes and more eco-friendly chemicals.Cellulose is the most abundant compound in spruce bark and has great potential to be used in various applications. The isolation of cellulose fibers from bark was in the present work carried out by first removing the extractives and non-cellulosic polysaccharides via sequential extractions using acetone and subcritical water. Nanocellulose was isolated from the bark cellulose and used to produce Pickering emulsions successfully. This proves that using side-stream products such as bark is feasible to produce high-value products like emulsions.Lignin is the second most abundant compound in the spruce bark. Following the acetone and subcritical water extractions, a mild cyclic organosolv extraction sequence was applied to be able to recover spruce bark lignin. The lignin extracted was comparable to lignin extracted from wood. Stilbene glucosides incorporated in the bark lignin provide the lignin with additional beneficial properties, i.e., antimicrobial and antioxidative. This elevates the value of the lignin further and makes it suitable for a variety of different applications.
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| 4. |
- Carvalho, Danila, 1984-
(författare)
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Study on the structure and properties of xylan extracted from eucalyptus, sugarcane bagasse and sugarcane straw
- 2015
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Lignocellulosic biomasses are an important source of chemical components such as cellulose, lignin and hemicelluloses, and can be used for a variety of purposes in both the pulp and paper and chemical conversion industries. Xylan, the main hemicellulose found in hardwood and grass plants, plays an important role during the pulping/pretreatment process reactions, including those used in 2nd generation bioethanol production. It may also play an important role in the production of certain novel materials.This thesis evaluates the composition of eucalyptus (Eucalyptus urophylla x Eucalyptus grandis), sugarcane bagasse and sugarcane straw, with a specific focus on the structure and properties of xylan. The chemical characterization of biomasses showed that sugarcane bagasse and straw contain larger amounts of extractives, ash and silica than eucalyptus. The large amount of silica leads to an overestimation of the Klason lignin content, if not corrected. By using a complete mass balance approach, sugarcane bagasse and straw were shown to contain smaller amounts of lignin (18.0% and 13.9%, respectively) than previously reported for these raw materials, and certainly a much smaller amount of lignin than was found in eucalyptus (27.4%). The hemicellulose content in sugarcane bagasse (28.7%) and straw (29.8%) was much higher than that in eucalyptus (20.3%).In order to investigate the structure of the xylan in greater detail, it was extracted with dimethyl sulfoxide from holocellulose, obtained by either peracetic acid or sodium chlorite delignification. The structure of the isolated xylans was confirmed by FTIR and 1H NMR analysis. In eucalyptus, the O-acetyl-(4-O-methylglucurono)xylan (MGX) was identified. This had a molar ratio of xylose units to branches of 4-O-methylglucuronic acid of 10:1.1 and a degree of acetylation of 0.39. All 4-O- methylglucuronic acid groups were attached to position O-2 of the xylose units, which had an acetyl group in position O-3. The acetyl groups were distributed in positions O-3 (64%), O-2 (26%) and O-2,3 (10%). The MGX had a molecular weight (Mw) of about 42 kDa.In bagasse and straw, arabinoxylan (AX) was identified. This had a molar ratio of xylose units to arabinosyl substitutions of 10:0.5 for bagasse and 10:0.6 for straw. A degree of acetylation was 0.29 and 0.08 for bagasse and straw, respectively. The arabinose units were attached preferentially to position O-3 in AX. In the xylan from bagasse, the acetyl groups were found in positions O-3 (60%), O-2 (13%) and O-2,3 (27%), while in the xylan from straw, the acetyl groups were distributed between positions O-3 (67%) and O-2 (33%). The AX had a molecular weight (Mw) of about 38 kDa and 30 kDa for bagasse and straw, respectively.The differences in the structure of xylan present in the various biomasses played an important role during hydrothermal pretreatment, which is often used as the first step in 2nd generation ethanol production. The varying amounts of uronic acid and acetyl groups resulted in different starting pH levels of liquor and, thus, affected the chemical transformation in the biomasses in different ways. The hydrothermal pretreatment resulted mostly in the removal and/or transformation of hemicelluloses, but also in the formation of a significant number of pseudo-lignin structures. In addition, in eucalyptus, pseudo-extractives structures were generated. The sugarcane straw showed the highest mass loss during the investigated pretreatment.
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| 5. |
- Ek, Patrik, 1980-
(författare)
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New methods for sensitive analysis with nanoelectrospray ionization mass spectrometry
- 2010
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In this thesis, new methods that address some current limitations in nanoelectrospray mass spectrometry (nESI-MS) analysis are presented. One of the major objectives is the potential gain in sensitivity that can be obtained when employing the proposed techniques. In the first part of this thesis, a new emitter, based on the generation of electrospray from a spray orifice with variable size, is presented. Electrospray is generated from an open gap between the edges of two individually mounted, pointed tips. The fabrication and evaluation of two different types of such emitters is presented; an ESI emitter fabricated from polyethylene terephtalate (Paper I), and a high-precision silicon device (Paper II). Both emitters were surface-treated in a selective way for an improved wetting of the gap and to confine the sample solution into the gap. In the second part of this thesis, different methods for improved sensitivity of nESI-MS analysis have been developed. In Paper III, a method for nESI-MS analysis from discrete sample volumes down to 1.5 nL is presented, using commercially available nESI needles. When analyzing attomole amounts of analyte in such a small volume of sample, an increased sensitivity was obtained, compared to when analyzing equal amounts in conventional nESI-MS analysis. To be able to analyze smaller sample volumes, needles with a narrower orifice and a higher flow resistance were needed. This triggered the development of a new method for fabrication of fused silica nESI needles (Paper IV). The fabrication is based on melting of a fused silica capillary by means of a rotating plasma, prior to pulling the capillary into a fine tip. Using the described technique, needles with sub-micrometer orifices could be fabricated. Such needles enabled the analysis of sample volumes down to 275 pL, and a further improvement of the sensitivity was obtained. In a final project (Paper V), nESI-MS was used to study the aggregation behavior of Aβ peptides, related to Alzheimer’s disease. An immunoprecipitation followed by nESI-MS was employed. This technique was also utilized to study the selectivity of the antibodies utilized.
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| 6. |
- Zhou, Yuye
(författare)
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Analysis of proteins related toautoimmune and inflammatorydiseases with focus on newenrichment methodologies and MALDI-TOF MS
- 2020
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Proteins have been widely studied in biological sample analysis due to the association with diseases. In the present project, method and technique development was carried out on the analysis of two proteins, immunoglobulin G (IgG) and osteopontin (OPN). The changes in IgG glycosylation, and elevated plasma OPN levels have been reported to be associated with chronic inflammation and autoimmune diseases.Lab synthesized zeolitic imidazolate framework (ZIF) nanocomposites (paper I) and environmentally friendly wood materials (paper II) were successfully utilized for IgG glycopeptide enrichment in order to eliminate the interference caused by non‑glycopeptides. The quantification and identification of glycopeptides by matrix assisted laser desorption/ionization - time of flight mass spectrometry (MALDI-TOF MS) was simplified using label-free internal standard, and intact glycopeptide identification without glycan release. The separation of enriched glycopeptides was further studied on capillary electrophoresis.Due to the low abundance of OPN in plasma, preconcentration of OPN is required prior to MS analysis. In paper III, a fast, cheap and antibody-free method was developed for recombinant human OPN (rhOPN) preconcentration from a complex mixture, human plasma, together with MALDI-TOF/TOF MS identification.In proteomics, enzymatic digestion of proteins is a very common step. In paper IV, the utilization of thiol-ene microchips (TE microchip) immobilized with trypsin provided fast digestion with residence time of only 10 s. Furthermore, the TE microchip linked with ascorbic acid could be used for IgG glycopeptide enrichment. These applications made TE microchips ready for multifunctional tasks in proteomics.Methods and techniques developed in the present project can be applied in the future for the study of biological samples, to investigate the possible relation between these proteins and autoimmune and inflammatory diseases, as well as other related diseases.
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| 7. |
- Bonn, Jonas, 1978-
(författare)
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Improved Techniques for Sampling and Sample Introduction in Gas Chromatography
- 2008
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Sampling and sample introduction are two key steps in quantitative gas chromatography. In this thesis, a development of a previously described sampling technique as well as a novel concept for sample introduction in gas chromatography are presented. The thesis is based on two papers. Paper I describes a method for preparing physically mixed polymers for use as sorbent phases in open tubular trapping of gaseous analytes. The concept is based on mechanical disintegration and mixing of solid or liquid poly(ethylene glycol), PEG, into poly(dimethylsiloxane), PDMS, in a straightforward manner. The resulting mixture exhibits a higher affinity towards polar analytes, as compared to pure PDMS. Paper II describes a novel approach to liquid sample introduction with the split/splitless inlet, used in gas chromatography. Classical injection techniques struggle with discrimination of high boiling analytes and poor repeatability of the injected amount of analytes. The presented injection technique utilizes high voltage to obtain a spraying effect of the injected liquid. The spraying effect can be achieved with a cold needle, which is unprecedented for gas chromatographic injections. The cold needle spraying results in highly repeatable injections, free from discrimination of high boiling analytes.
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| 8. |
- Jacksén, Johan, 1980-
(författare)
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Improved techniques for CE and MALDI-MS including microfluidic hyphenations foranalysis of biomolecules
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- In this thesis, improved techniques for biomolecule analysis using capillary electrophoresis (CE) and matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) and hyphenations between those have been presented.A pre-concentration method which is possible to apply in both techniques, has also been investigated. In this work the off-line MS mode has been used either in the form of fractionation (Paper I) or by incorporating the MALDI target in the CE separation system (Paper II).In Paper I, a protocol for CE-MALDI analysis of cyanogen bromide digested bacteriorhodopsin (BR) peptides as model integral membrane protein peptides were established. Also, an improved protocol for partially automated manufacturing of a concentration MALDI-target plate is presented. The design of the targets was suitable for the fractions from the CE. A novel technique for the integration of CE to MALDI-MS using a closed-open-closed system is presented in Paper II, where the open part is a micro canal functioning as a MALDI target window. A protein separation was obtained and detected with MALDI-MS analysis in the micro canal. A method has been developed for detection of monosaccharides originating from hydrolysis of a single wood fiber performed in a micro channel, with an incorporated electromigration pre-concentration step preceding CE analysis in Paper III. The pre-concentration showed to be highly complex due to the fact that several parameters are included that affecting each other. In Paper IV a protocol using enzymatic digestion, MALDI-TOF-MS and CE with laser induced fluorescence (LIF) detection for the investigation of the degree of substitution of fluorescein isothiocyanate (FITC) to bovine serum albumin (BSA), as a contact allergen model system for protein-hapten binding in the skin, is presented. The intention of a further CE-MALDI hyphenation has been considered during the work. In Paper V 2,6-dihydroxyacetophenone (DHAP) was investigated, showing promising MALDI-MS matrix properties for hydrophobic proteins and peptides. 2,5-dihydroxybenzoic acid (DHB) was undoubtedly the better matrix for the hydrophilic proteins, but its performance for the larger and hydrophobic peptides was not optimal. Consequently, DHAP can be used as a compliment matrix for improved analysis of hydrophobic analytes.
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| 9. |
- Jacksén, Johan, 1980-
(författare)
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Improved techniques for CE-MALDI-MS off-line coupling and MALDI-MS analysis of primarily hydrophobic proteins and peptides
- 2007
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Due to the hydrophobic nature of integral membrane proteins (IMP) they give rise to several difficulties concerning handling and analysis, which is not the case for the most water soluble proteins. New analysis methods are needed, where the insolubility problems of the hydrophobic proteins due to aggregation and adhesion are tackled. Those problems also affect digestion performance and equipment compatibility for the analysis. Protocols for analysis and separation specified for IMP are presented in Paper I and III. The instrumentation used in this work was capillary electrophoresis (CE) and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). Both instruments are suitable for peptide/proteins analysis. In Paper I, protocols for a CE separation of bacteriorhodopsin (BR) peptides as model IMP peptides are established. Also, a partially automated manufacturing procedure of a concentration MALDI-target is presented, suitable for fractions from CE. The MS analysis detected 9 out of 10 cyanogen bromide (CNBr) digested BR peptides. A novel technique for the off-line integration of CE to MALDI-MS using a closed-open-closed system is presented in Paper II, where the open part is a microcanal functioning as a MALDI target window. Investigation of the microcanal electro-osmotic flow (EOF) properties and band broadening characteristics was performed. A protein separation was obtained and detected with MALDI-MS analysis in the microcanal. Different protein digestion methods were evaluated using BR in Paper III through MALDI-MS. Several digestion methods as well as MS media were investigated alongside different MALDI matrices. For example, matrices as the hydrophobic 2,6-dihydroxyacetophenone (DHAP) and 2-Hydroxy-3-methoxybenzoic acid (2H3MBA) or 2-Hydroxy-5-methoxybenzoic acid (2H5MBA) mixed with DHB, appeared to be promising matrices for analysis of BR.
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| 10. |
- Josefsson, Leila
(författare)
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Bioanalysis using capillary electrophoresis and mass spectrometry : Applied on proteins, protein nanofibrils and polyvinyl alcohol microbubbles
- 2019
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The sequencing of the genome of various species, including the human species, have led to increased understanding about how a protein structure is generated, and how specific structures are related to the proteins’ functionality. In paper I and II of this thesis, the folding of proteins in vitro to form hierarchical nanostructures, which in vivo often have a pathological effect, have been studied. Protein isolates from soybean and potato, that are byproducts from oil and starch production, respectively, were used as a starting material for protein nanofibril (PNF) formation, and mass spectrometry was used to identify the building blocks that are included in the formed PNF. The five peptides identified in soybean PNF and the six peptides identified in potato PNF originated from the major seed storage proteins for the respective crop.The use of ionic liquids has increased for improvement of the performance of different separation techniques due to their adjustable properties, and good solvating ability. In paper III, an ionic liquid and water mixture was used as background electrolyte in capillary electrophoresis for protein separation. The system showed high reproducibility at basic conditions, and could potentially be used for routine control analysis.Many diseases and injuries require clinical diagnosis techniques e.g. ultrasound imaging, to be detected, and for the physician to be able to decide the correct therapy. To increase the resolution of such imaging techniques, contrast agents can be used. In paper IV-VI, a newly developed contrast agent consisting of air-filled microbubbles stabilized with a shell of polyvinyl alcohol (PVA-MBs) was studied. Development of a capillary electrophoretic method for analysis of the PVA-MBs with the intentions to be used for clinical diagnosis is performed, where different detectors such as a UV detector, a UV area imaging detector and an in-house constructed microscope are used to increase the sensitivity of detection for the PVA-MBs. The developed method could be used for quantification of the contrast agent, since individual PVA-MBs were visible using the imaging detectors. Findings regarding the mobility of the PVA-MBs in human blood plasma and in water implies that a protein corona was formed around the MBs.
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