Asymmetrical flow field-flow fractionation in the study of water-soluble macromolecules

Asymmetrical flow field-flow fractionation (AsFlFFF) was constructed, and its applicability to industrial, biochemical, and pharmaceutical applications was studied. The effect of several parameters, such as pH, ionic strength, temperature and the reactants mixing ratios on the particle sizes, molar masses, and the formation of aggregates of macromolecules was determined by AsFlFFF. In the case of industrial application AsFlFFF proved to be a valuable tool in the characterization of the hydrodynamic particle sizes, molar masses and phase transition behavior of various poly(N-isopropylacrylamide) (PNIPAM) polymers as a function of viscosity and phase transition temperatures. The effect of sodium chloride salt and the molar ratio of cationic and anionic polyelectrolytes on the hydrodynamic particle sizes of poly (methacryloxyethyl trimethylammonium chloride) and poly (ethylene oxide)-block-poly (sodium methacrylate) and their complexes were studied. The particle sizes of PNIPAM polymers, and polyelectrolyte complexes measured by AsFlFFF were in agreement with those obtained by dynamic light scattering. The molar masses of PNIPAM polymers obtained by AsFlFFF and size exclusion chromatography agreed also well. In addition, AsFlFFF proved to be a practical technique in thermo responsive behavior studies of polymers at temperatures up to about 50 oC. The suitability of AsFlFFF for biological, biomedical, and pharmaceutical applications was proved, upon studying the lipid-protein/peptide interactions, and the stability of liposomes at different temperatures. AsFlFFF was applied to the studies on the hydrophobic and electrostatic interactions between cytochrome c (a basic peripheral protein) and anionic lipid, and oleic acid, and sodium dodecyl sulphate surfactant. A miniaturized AsFlFFF constructed in this study was exploited in the elucidation of the effect of copper (II), pH, ionic strength, and vortexing on the particle sizes of low-density lipoproteins.

Late Transition Metal and Aluminum Complexes for the Polymerization of Ethene and Acrylates

Polyethene, polyacrylates and polymethyl acrylates are versatile materials that find wide variety of applications in several areas. Therefore, polymerization of ethene, acrylates and methacrylates has achieved a lot attention during past years. Numbers of metal catalysts have been introduced in order to control the polymerization and to produce tailored polymer structures. Herein an overview on the possible polymerization pathways for ethene, acrylates and methacrylates is presented. In this thesis iron(II) and cobalt(II) complexes bearing tri- and tetradentate nitrogen ligands were synthesized and studied in the polymerization of tertbutyl acrylate (tBA) and methyl methacrylate (MMA). Complexes are activated with methylaluminoxane (MAO) before they form active combinations for polymerization reactions. The effect of reaction conditions, i.e. monomer concentration, reaction time, temperature, MAO to metal ratio, on activity and polymer properties were investigated. The described polymerization system enables mild reaction conditions, the possibility to tailor molar mass of the produced polymers and provides good control over the polymerization. Moreover, the polymerization of MMA in the presence of iron(II) complex with tetradentate nitrogen ligands under conditions of atom transfer radical polymerization (ATRP) was studied. Several manganese(II) complexes were studied in the ethene polymerization with combinatorial methods and new active catalysts were found. These complexes were also studied in acrylate and methacrylate polymerizations after MAO activation and converted into the corresponding alkyl (methyl or benzyl) derivatives. Combinatorial methods were introduced to discover aluminum alkyl complexes for the polymerization of acrylates and methacrylates. Various combinations of aluminum alkyls and ligands, including phosphines, salicylaldimines and nitrogen donor ligands, were prepared in situ and utilized to initiate the polymerization of tBA. Phosphine ligands were found to be the most active and the polymerization MMA was studied with these active combinations. In addition, a plausible polymerization mechanism for MMA based on ESI-MS, 1H and 13C NMR is proposed.

Stabilization of Phospholipid Coatings in Capillary Electrophoresis

The development of a simple method of coating a semi-permanent phospholipid layer onto a capillary for electrochromatography use was the focus of this study. The work involved finding good coating conditions, stabilizing the phospholipid coating, and examining the effect of adding divalent cations, cetyltrimethylammonium bromide, and polyethylene glycol (PEG)-lipids on the stability of the coating. Since a further purpose was to move toward more biological membrane coatings, the capillaries were also coated with cholesterol-containing liposomes and liposomes of red blood cell ghost lipids. Liposomes were prepared by extrusion, and large unilamellar vesicles with a diameter of about 100 nm were obtained. Zwitterionic phosphatidylcholine (PC) was used as a basic component, mainly 1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine (POPC) but also eggPC and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). Different amounts of sphingomyelin, bovine brain phosphatidylserine, and cholesterol were added to the PC. The stability of the coating in 40 mM N-(2-hydroxyethyl)piperazine-N’-(2-ethanesulfonic acid) (HEPES) solution at pH 7.4 was studied by measuring the electroosmotic flow and by separating neutral steroids, basic proteins, and low-molar-mass drugs. The presence of PC in the coating solution was found to be essential to achieving a coating. The stability of the coating was improved by the addition of negative phosphatidylserine, cholesterol, divalent cations, or PEGylated lipids, and by working in the gel-state region of the phospholipid. Study of the effect on the PC coating of divalent metal ions calcium, magnesium, and zinc showed a molar ratio of 1:3 PC/Ca2+ or PC/Mg2+ to give increased rigidity to the membrane and the best coating stability. The PEGylated lipids used in the study were sterically stabilized commercial lipids with covalently attached PEG chains. The vesicle size generally decreased when PEGylated lipids of higher molar mass were present in the vesicle. The predominance of discoidal micelles over liposomes increased PEG chain length and the average size of the vesicles thus decreased. In the capillary electrophoresis (CE) measurements a highly stable electroosmotic flow was achieved with 20% PEGylated lipid in the POPC coating dispersion, the best results being obtained for disteroyl PEG (3000) conjugates. The results suggest that smaller particles (discoidal micelles) result in tighter packing and better shielding of silanol groups on the silica wall. The effect of temperature on the coating stability was investigated by using DPPC liposomes at temperatures above (45 C) and below (25 C) the main phase transition temperature. Better results were obtained with DPPC in the more rigid gel state than in the fluid state: the electroosmotic flow was heavily suppressed and the PC coating was stabilized. Also dispersions of DPPC with 0−30 mol% of cholesterol and sphingomyelin in different ratios, which more closely resemble natural membranes, resulted in stable coatings. Finally, the CE measurements revealed that a stable coating is formed when capillaries are coated with liposomes of red blood cell ghost lipids.

Selective-Area Atomic Layer Deposition

Atomic layer deposition (ALD) is a method to deposit thin films from gaseous precursors to the substrate layer-by-layer so that the film thickness can be tailored with atomic layer accuracy. Film tailoring is even further emphasized with selective-area ALD which enables the film growth to be controlled also on the substrate surface. Selective-area ALD allows the decrease of a process steps in preparing thin film devices. This can be of a great technological importance when the ALD films become into wider use in different applications. Selective-area ALD can be achieved by passivation or activation of a surface. In this work ALD growth was prevented by octadecyltrimethoxysilane, octadecyltrichlorosilane and 1-dodecanethiol SAMs, and by PMMA (polymethyl methacrylate) and PVP (poly(vinyl pyrrolidone) polymer films. SAMs were prepared from vapor phase and by microcontact printing, and polymer films were spin coated. Microcontact printing created patterned SAMs at once. The SAMs prepared from vapor phase and the polymer mask layers were patterned by UV lithography or lift-off process so that after preparation of a continuous mask layer selected areas of them were removed. On these areas the ALD film was deposited selectively. SAMs and polymer films prevented the growth in several ALD processes such as iridium, ruthenium, platinum, TiO2 and polyimide so that the ALD films did grow only on areas without SAM or polymer mask layer. PMMA and PVP films also protected the surface against Al2O3 and ZrO2 growth. Activation of the surface for ALD of ruthenium was achieved by preparing a RuOX layer by microcontact printing. At low temperatures the RuCp2-O2 process nucleated only on this oxidative activation layer but not on bare silicon.

Desorptio/fotoionisaatio ilmanpaineessa takavarikoitujen huumausaineiden, steroidien ja räjähdysaineiden tutkimuksessa

Tutkimuksen tarkoituksena oli selvittää desorptio/fotoionisaatio ilmanpaineessa tekniikan (engl. desorption atmospheric pressure photoionization, DAPPI) soveltuvuutta rikosteknisen laboratorion näytteiden analysointiin. DAPPI on nopea massaspektrometrinen ionisaatiotekniikka, jolla voidaan tutkia yhdisteitä suoraan erilaisilta pinnoilta. DAPPI:ssa käytetään lämmitettyä mikrosirua, joka suihkuttaa höyrystynyttä liuotin- ja kaasuvirtausta kohti näytettä. Näytteen pinnan komponentit desorboituvat lämmön vaikutuksesta, jonka jälkeen ionisoituminen tapahtuu VUV-lampun emittoimien fotonien avulla.DAPPI:lla tutkittiin takavarikoituja huumausaineita, anabolisia steroideja ja räjähdysaineita sekä niiden jäämiä erilaisilta pinnoilta. Lisäksi kartoitettiin DAPPI:n mahdollisuuksia ja rajoituksia erilaisille näytematriiseille ilman näytteiden esikäsittelyä. Takavarikoitujen huumausaineiden tutkimuksessa analysoitiin erilaisia tabletteja, jauheita, kasvirouheita, huumekasveja (khat, oopium, kannabis) ja sieniä. Anabolisia steroideja tunnistettiin tableteista sekä ampulleista, jotka sisälsivät öljymäistä nestettä. Jauheet ripoteltiin kaksipuoliselle teipille ja analysoitiin siltä. Muut näytteet analysoitiin sellaisenaan ilman minkäänlaista esikäsittelyä, paitsi nestemäisten näytteiden kohdalla näyte pipetoitiin talouspaperille, joka analysoitiin DAPPI:lla. DAPPI osoittautui nopeaksi ja yksinkertaiseksi menetelmäksi takavarikoitujen huumausaineiden ja steroidien analysoimisessa. Se soveltui hyvin rikoslaboratorion erityyppisten näytteiden rutiiniseulontaan ja helpotti erityisesti huumekasvien ja öljymäisten steroidiliuosten tutkimusta. Massaspektrometrin likaantuminen pystyttiin ehkäisemään säätämällä näytteen etäisyyttä sen suuaukosta. Likaantumista ei havaittu huolimatta näytteiden korkeista konsentraatioista ja useita kuukausia jatkuneista mittauksista. Räjähdysaineiden tutkimuksessa keskityttiin seitsemän eri räjähdysaineen DAPPI-MS-menetelmän kehitykseen; trinitrotolueeni (TNT), nitroglykoli (NK), nitroglyseriini (NG), pentriitti (PETN), heksogeeni (RDX), oktogeeni (HMX) ja pikriinihappoä Nämä orgaaniset räjähteet ovat nitraattiyhdisteitä, jotka voidaan jakaa rakenteen puolesta nitroamiineihin (RDX ja HMX), nitroaromaatteihin (TNT ja pikriinihappo) sekä nitraattiestereihin (PETN, NG ja NK). Menetelmäkehityksessä räjähdysainelaimennokset pipetoitiin polymetyylimetakrylaatin (PMMA) päälle ja analysoitiin siitä. DAPPI:lla tutkittiin myäs autenttisia räjähdysainejäämiä erilaisista matriiseista. DAPPI:lla optimoitiin jokaiselle räjähdysaineelle sopiva menetelmä ja yhdisteet saatiin näkymään puhdasaineina. Räjähdysainejäämien analysoiminen erilaisista rikospaikkamateriaaleista osoittautui haastavammaksi tehtäväksi, koska matriisit aiheuttivat itsessään korkean taustan spektriin, josta räjähdysaineiden piikit eivät useimmiten erottuneet tarpeeksi. Muut desorptioionisaatiotekniikat saattavat soveltua paremmin haastavien räjähdysainejäämien havaitsemiseksi.