Investigation of single tryptophan proteins encapsulated in TEOS-derived sol-gel matrices by fluorescence spectroscopy /

In the work reported here, optically clear, ultrathin TEOS derived sol-gel slides which were suitable for studies of tryptophan (Trp) fluorescence from entrapped proteins were prepared by the sol-gel technique and characterized. The monitoring of intrinsic protein fluorescence provided information about the structure and environment of the entrapped protein, and about the kinetics of the interaction between the entrapped protein and extemal reagents. Initial studies concentrated on the single Trp protein monellin which was entrapped into the sol-gel matrices. Two types of sol-gel slides, termed "wet aged", in which the gels were aged in buffer and "dry-aged", in which the gels were aged in air , were studied in order to compare the effect of the sol-gel matrix on the structure of the protein at different aging stages. Fluorescence results suggested that the mobility of solvent inside the slides was substantially reduced. The interaction of the entrapped protein with both neutral and charged species was examined and indicated response times on the order of minutes. In the case of the neutral species the kinetics were diffusion limited in solution, but were best described by a sum of first order rate constants when the reactions occurred in the glass matrix. For charged species, interactions between the analytes and the negatively charged glass matrix caused the reaction kinetics to become complex, with the overall reaction rate depending on both the type of aging and the charge on the analyte. The stability and conformational flexibility of the entrapped monellin were also studied. These studies indicated that the encapsulation of monellin into dry-aged monoliths caused the thermal unfolding transition to broaden and shift upward by 14°C, and causedthe long-term stability to improve by 12-fold (compared to solution). Chemical stability studies also showed a broader transition for the unfolding of the protein in dry-aged monoliths, and suggested that the protein was present in a distribution of environments. Results indicated that the entrapped proteins had a smaller range of conformational motions compared to proteins in solution, and that entrapped proteins were not able to unfold completely. The restriction of conformational motion, along with the increased structural order of the internal environment of the gels, likely resulted in the improvements in themial and long-term stability that were observed. A second protein which was also studied in this work is the metal binding protein rat oncomodulin. Initially, the unfolding behavior of this protein in aqueous solution was examined. Several single tryptophan mutants of the metal-binding protein rat oncomodulin (OM) were examined; F102W, Y57W, Y65W and the engineered protein CDOM33 which had all 12 residues of the CD loop replaced with a higher affinity binding loop. Both the thermal and the chemical stability were improved upon binding of metal ions with the order apo < Ca^^ < Tb^"^. During thermal denaturation, the transition midpoints (Tun) of Y65W appeared to be the lowest, followed by Y57W and F102W. The placement of the Trp residue in the F-helix in F102W apparently made the protein slightly more thermostable, although the fluorescence response was readily affected by chemical denaturants, which probably acted through the disruption of hydrogen bonds at the Cterminal end of the F-helix. Under both thermal and chemical denaturation, the engineered protein showed the highest stability. This indicated that increasing the number of metal ligating oxygens in the binding site, either by using a metal ion with a higher coordinatenumber (i.e. Tb^*) which binds more carboxylate ligands, or by providing more ligating groups, as in the CDOM33 replacement, produces notable improvements in protein stability. Y57W and CE)OM33 OM were chosen for further studies when encapsulated into sol-gel derived matrices. The kinetics of interaction of terbium with the entrapped proteins, the ability of the entrapped protein to binding terbium, as well as thermal stability of these two entrapped protein were compared with different levels of Ca^"*^ present in the matrix and in solution. Results suggested that for both of the proteins, the response time and the ability to bind terbium could be adjusted by adding excess calcium to the matrix before gelation. However, the less stable protein Y57W only retained at most 45% of its binding ability in solution while the more stable protein CDOM33 was able to retain 100% binding ability. Themially induced denaturation also suggested that CDOM33 showed similar stability to the protein in solution while Y57W was destabilized. All these results suggested that "hard" proteins (i.e. very stable) can easily survive the sol-gel encapsulation process, but "soft" proteins with lower thermodynamic stability may not be able to withstand the sol-gel process. However, it is possible to control many parameters in order to successfully entrap biological molecules into the sol-gel matrices with maxunum retention of activity.

Melting the matrices : structure, anti-structure, and the emerging conversation /

This thesis investigates two cases of Christian churches, which as a part of their mission seek to accommodate people who would otherwise not be interested in church. One of these communities consider themselves a part of the global 'emerging church' movement, and the other does not. I argue that both communities are employing what I call 'de-compartmentalization' strategy in order to adopt a pragmatic relationship with social and political issues. Furthermore I discuss the case of the emerging church community as an example of 'paraliminal community '; a concept I develop from the work of Victor Turner and Arnold van Gennep.

Fluorometric monitoring of the properties of proteins entrapped in sol-gel derived matrices

The successful development of stable biosensors incorporating entrapped proteins suffers from poor understanding of the properties of the entrapped biomolecules. This thesis reports on the use of fluorescence spectroscopy to investigate the properties of proteins entrapped in sol-gel processed silicate materials. Two different single tryptophan (Trp) proteins were investigated in this thesis, the Ca2 + binding protein cod III parvalbumin (C3P) and the salicylate binding protein human serum albumin (HSA). Furthermore, the reactive single cysteine (Cys) residue within C3P and HSA were labelled with the probes iodoacetoxynitrobenzoxadiazole (C3P) and acrylodan (C3P and HSA) to further examine the structure, stability and function of the free and entrapped proteins. The results show that both C3P and HSA can be successfully entrapped into sol-gelderived matrices with retention of function and conformational flexibility.

Predicting performance on fluid intelligence from speed of processing, working memory, and controlled attention

Fluid inteliigence has been defined as an innate ability to reason which is measured commonly by the Raven's Progressive Matrices (RPM). Individual differences in fluid intelligence are currently explained by the Cascade model (Fry & Hale, 1996) and the Controlled Attention hypothesis (Engle, Kane, & Tuholski, 1999; Kane & Engle, 2002). The first theory is based on a complex relation among age, speed, and working memory which is described as a Cascade. The alternative to this theory, the Controlled Attention hypothesis, is based on the proposition that it is the executive attention component of working memory that explains performance on fluid intelligence tests. The first goal of this study was to examine whether the Cascade model is consistent within the visuo-spatial and verbal-numerical modalities. The second goal was to examine whether the executive attention component ofworking memory accounts for the relation between working memory and fluid intelligence. Two hundred and six undergraduate students between the ages of 18 and 28 completed a battery of cognitive tests selected to measure processing speed, working memory, and controlled attention which were selected from two cognitive modalities, verbalnumerical and visuo-spatial. These were used to predict performance on two standard measures of fluid intelligence: the Raven's Progressive Matrices (RPM) and the Shipley Institute of Living Scales (SILS) subtests. Multiple regression and Structural Equation Modeling (SEM) were used to test the Cascade model and to determine the independent and joint effects of controlled attention and working memory on general fluid intelligence. Among the processing speed measures only spatial scan was related to the RPM. No other significant relations were observed between processing speed and fluid intelligence. As 1 a construct, working memory was related to the fluid intelligence tests. Consistent with the predictions for the RPM there was support for the Cascade model within the visuo-spatial modality but not within the verbal-numerical modality. There was no support for the Cascade model with respect to the SILS tests. SEM revealed that there was a direct path between controlled attention and RPM and between working memory and RPM. However, a significant path between set switching and RPM explained the relation between controlled attention and RPM. The prediction that controlled attention mediated the relation between working memory and RPM was therefore not supported. The findings support the view that the Cascade model may not adequately explain individual differences in fluid intelligence and this may be due to the differential relations observed between working memory and fluid intelligence across different modalities. The findings also show that working memory is not a domain-general construct and as a result its relation with fluid intelligence may be dependent on the nature of the working memory modality.

Phonon spectra and thermal properties of some fcc metals using embedded-atom potentials /

By employing the embedded-atom potentials of Mei et ai.[l], we have calculated the dynamical matrices and phonon dispersion curves for six fee metals (Cu,Ag,Au,Ni,Pd and Pt). We have also investigated, within the quasiharmonic approximation, some other thermal properties of these metals which depend on the phonon density of states, such as the temperature dependence of lattice constant, coefficient of linear thermal expansion, isothermal and adiabatic bulk moduli, heat capacities at constant volume and constant pressure, Griineisen parameter and Debye temperature. The computed results are compared with the experimental findings wherever possible. The comparison shows a generally good agreement between the theoretical values and experimental data for all properties except the discrepancies of phonon frequencies and Debye temperature for Pd, Pt and Au. Further, we modify the parameters of this model for Pd and Pt and obtain the phonon dispersion curves which is in good agreement with experimental data.

Exchange energy calculation of He-He with supermolecular one-matrix

Exch~nge energy of the He-He system is calculated using the one-density matrix which has been modified according to the supermolecular density formula quoted by Kolos. The exchange energy integrals are computed analytically and by the Monte Carlo method. The results obtained from both ways compared favourably,with the results obtained from the SCF program HONDO

Microstructural and geochemical investigation of North American lower-middle Paleozoic trilobites and recent arthropods

Trilobites ¥tere collected from Ordovician and Devonian formations of Ontario} New York} Ohio} Oklahoma} and Indiana. Diversity was generally low} but 19..?telllS and Ph..~tY>ps ¥tere the most abundant species from the Ordovician and Devonian} respectively. Recent marine arthropods ¥tere collected from the Atlantic shore of the middle Florida Keys} and from the Pacific and lagoonal waters at Cape Beale} B. C. Fresh-water arthropods were collected along the shore of the Severn River in northcentral Ontario. Cuticles ¥tere analyzed for major} minor and trace elements, 180 and 13C isotopes, as ¥tell as examined by scanning electron micr?scope to identify original and diagenetic fabrics. Examination of trilobite cuticles by scanning electron microscope revealed several microstructures consistent with those observed in Recent arthropods. Microstructures} such as setae and tegumental gland duct openings} in like sized Lim/IllS and Isoteline trilobites may indicate common ancestral origins for these organisms, or simply parallel cuticle evolutions. The dendritic microstructure, originally' thought to be a diagenetic indicator, was found in Recent specimens and therefore its presence in trilobites may be suggestive of the delicate nature of diagenesis in trilobites. The absence of other primary microstructures in trilobites may indicate alteration, taxonomic control} or that there is some inherent feature of S EM examination which may' not allow detection of some features} while others are apparently visit·le onl~1 under SH.·1. The region of the cuticle sampled for examination is also a major influence in detecting pristine microstructures, as not all areas of trilobite and Recent arthropod cuticles will have microstructures identifiable in a SEM study. Subtleties in the process of alteration, however} ma~·· leave pristine microstructures in cuticles that are partial~/ silicified or do 10m itized, and degree and type of alteration may vary stratigraphically and longitudinally within a unit. The presence of fused matrices, angular calcite rhombs, and pyrite in the cuticle are thought to be indicative of altered cuticles, although pyritization may not affect the entire cuticle. t-~atural processes in Recent arthropods, such as molting, lead to variations in cuticle chemistries, and are thought to reflect the area of concentration of the elements during calcification. The level of sodium in Recent arthropods was found to be higher than that in trilobites, but highly mobile when sUbjected to the actions of VY'€'athering. Less saline water produced lovy'€'r magnesium and higher calcium values in Recent specimens .. and metal variations in pristine Ordovician trilobite cuticle appears to follow the constraints outlined for Recent arthropods, of regulation due to the chemislry of the surrounding medium. In diagenetic analysis, sodium, strontium and magnesium proved most beneficial in separating altered from least altered trilobites. Using this criterion, specimens from shale show the least amount of geochemical alteration, and have an original mineralogy of 1.7 - 2.4 mole % MgC03 (8000 t(> 9500 ppm magnesium) for both /s>..?/e/11S lJA'i.riff!11S and PseIAit'11J17ites I..itmirpin..itl/~ and 2.8 - 3.3 mole % MgC03 (5000 to 7000 ppm magnesium) for Ph.i{).?PS This is Slightly lower than the mineralogy of Recent marine arthropods (4.43 - 12.1 mole % MgC03), and slightly higher than that of fresh-water crayfish (0.96 - 1.82 mole % MgC03). Geochemically pristine trilobites were also found to possess primary microstructures. Stable isotope values and trends support the assertion that marine-meteoriclburial fluids were responsible for the alteration observed in a number of the trilobite specimens. The results of this stUdy suggest that fossil material has to be evaluated separately along taxonomic and lithological lines to arrive at sensible diagenetic and e nvironmenta I interpretations.

Gas chromatographic electron capture negative ionization mass spectrometric (GC/ECNI/MS) determination of unique fluorinated compounds in the sediments of Lake Ontario and the effect of high-boiling alcohols (as injection solvents) on chromatographic behaviour of polycyclic aromatic hydrocarbons in gas chromatography

Part I - Fluorinated Compounds A method has been developed for the extraction, concentration, and determination of two unique fluorinated compounds from the sediments of Lake Ontario. These compounds originated from a common industrial landfill, and have been carried to Lake Ontario by the Niagara River. Sediment samples from the Mississauga basin of Lake Ontario have been evaluated for these compounds and a depositional trend was established. The sediments were extracted by accelerated solvent extraction (ASE) and then underwent clean-up, fractionation, solvent exchange, and were concentrated by reduction under nitrogen gas. The concentrated extracts were analyzed by gas chromatography - electron capture negative ionization - mass spectrometry. The depositional profile determined here is reflective of the operation of the landfill and shows that these compounds are still found at concentrations well above background levels. These increased levels have been attributed to physical disturbances of previously deposited contaminated sediments, and probable continued leaching from the dumpsite. Part II - Polycyclic Aromatic Hydrocarbons Gas chromatography/mass spectrometry is the most common method for the determination of polycyclic aromatic hydrocarbons (PAHs) from various matrices. Mass discrimination of high-boiling compounds in gas chromatographic methods is well known. The use of high-boiling injection solvents shows substantial increase in the response of late-eluting peaks. These solvents have an increased efficiently in the transfer of solutes from the injector to the analytical column. The effect of I-butanol, I-pentanol, cyclopentanol, I-hexanol, toluene and n-octane, as injection solvents, was studied. Higher-boiling solvents yield increased response for all PAHs. I -Hexanol is the best solvent, in terms of P AH response, but in this solvent P AHs were more susceptible to chromatographic problems such as peak splitting and tailing. Toluene was found to be the most forgiving solvent in terms of peak symmetry and response. It offered the smallest discrepancies in response, and symmetry over a wide range of initial column temperatures.

ReAlM - a system to manipulate relations

Given a heterogeneous relation algebra R, it is well known that the algebra of matrices with coefficient from R is relation algebra with relational sums that is not necessarily finite. When a relational product exists or the point axiom is given, we can represent the relation algebra by concrete binary relations between sets, which means the algebra may be seen as an algebra of Boolean matrices. However, it is not possible to represent every relation algebra. It is well known that the smallest relation algebra that is not representable has only 16 elements. Such an algebra can not be put in a Boolean matrix form.[15] In [15, 16] it was shown that every relation algebra R with relational sums and sub-objects is equivalent to an algebra of matrices over a suitable basis. This basis is given by the integral objects of R, and is, compared to R, much smaller. Aim of my thesis is to develop a system called ReAlM - Relation Algebra Manipulator - that is capable of visualizing computations in arbitrary relation algebras using the matrix approach.

Characterisation and optimisation of the flavour of health-promoting, plantderived bitterants in functional beverages.

Flavour is a combination of taste, odour, and chemesthetic sensations. Close associations exist between these sensory modalities, and thus, the overall flavour of a food or beverage product can change when the intensity of one or more of these sensations is altered. Strategies to modify flavour are often utilized by the food industry, and are central to the engineering of new and reformulated products. For functional food and beverages, flavour modification is particularly important, as fortifying agents can elicit high levels of less than desirable sensations, such as bitterness and astringency. The application of various flavour modifying strategies can decrease the perceived intensity of these sensations, and in tum, improve the sensory profile of the product. This collection of studies describes the sensory characteristics of experimental functional beverages fortified with trans-resveratrol, (+)-catechin, and/or caffeine, and examines the impact of novel flavour modifying strategies on the perceived flavour of these beverages. In the first study, results demonstrate that the flavour profile of Cabemet Sauvignon wines fortified with 20 mglL and 200 mg/L of trans-resveratrol is not perceived as different compared to control wine (0 mglL). However, Riesling wine fortified with 200 mg/L is perceived as significantly higher in bitterness compared to 20 mglL and control. For some functional food formulations, alternative strategies for flavour modification are needed. Traditional methods, such as the addition of sucrose and sodium chloride, may decrease the perceived 'healthiness' of a product, and thus, may be sub-optimal. In a second study, high and low concentrations of five different bitter inhibiting compounds - 'bitter blockers' - (B-cyclodextrin, homoeridictyol sodium salt, carboxymethylcellulose - low viscosity, zinc sulfate, magnesium sulfate) were tested for their efficacy towards decreasing the bitterness of high and low concentrations of caffeine and (+)catechin - two health-relevant, plant-derived bitterants. B-cyclodextrin and homoeridictyol sodium salt were the most effective blockers at decreasing (+ )-catechin and caffeine, respectively. In addition to bitter blockers, additional flavour modifying strategies, either alone or in combination - may also be successful in functional food formulations. Both sucrose and rebaudioside A - a plant-derived sweetener - were effective at decreasing the bitterness of (+)catechin. When added to (+)-catechin along with B-cyc1odextrin, both sweeteners provided the most effective decrease in bitterness compared to binary, ternary, or quaternary mixtures of (+)catechin together with bitter blockers, sweeteners, andlor odourants. The perceived intensity of sensations elicited by sweeteners and odourants was not affected by the addition of bitter blockers, and thus, their impact within these complex matrices is minimal. In addition, withinmodal (taste-taste) compared to cross-modal (taste-odour) sensory interactions were more effective at decreasing the bitterness of (+ )-catechin. Overall, results from these studies demonstrate that certain novel, alternative flavour modifying approaches may be successful towards lowering the bitterness and astringency elicited by (+ )-catechin and caffeine in aqueous solutions.

RelMDD-A Library for Manipulating Relations Based on MDDs

Relation algebras is one of the state-of-the-art means used by mathematicians and computer scientists for solving very complex problems. As a result, a computer algebra system for relation algebras called RelView has been developed at Kiel University. RelView works within the standard model of relation algebras. On the other hand, relation algebras do have other models which may have different properties. For example, in the standard model we always have L;L=L (the composition of two (heterogeneous) universal relations yields a universal relation). This is not true in some non-standard models. Therefore, any example in RelView will always satisfy this property even though it is not true in general. On the other hand, it has been shown that every relation algebra with relational sums and subobjects can be seen as matrix algebra similar to the correspondence of binary relations between sets and Boolean matrices. The aim of my research is to develop a new system that works with both standard and non-standard models for arbitrary relations using multiple-valued decision diagrams (MDDs). This system will implement relations as matrix algebras. The proposed structure is a library written in C which can be imported by other languages such as Java or Haskell.

Application of thermal ion-molecule reactions in tackling spectroscopic inferences in inductively coupled plasma mass spectrometry

Part I: Ultra-trace determination of vanadium in lake sediments: a performance comparison using O2, N20, and NH3 as reaction gases in ICP-DRC-MS Thermal ion-molecule reactions, targeting removal of specific spectroscopic interference problems, have become a powerful tool for method development in quadrupole based inductively coupled plasma mass spectrometry (ICP-MS) applications. A study was conducted to develop an accurate method for the determination of vanadium in lake sediment samples by ICP-MS, coupled with a dynamic reaction cell (DRC), using two differenvchemical resolution strategies: a) direct removal of interfering C10+ and b) vanadium oxidation to VO+. The performance of three reaction gases that are suitable for handling vanadium interference in the dynamic reaction cell was systematically studied and evaluated: ammonia for C10+ removal and oxygen and nitrous oxide for oxidation. Although it was able to produce comparable results for vanadium to those using oxygen and nitrous oxide, NH3 did not completely eliminate a matrix effect, caused by the presence of chloride, and required large scale dilutions (and a concomitant increase in variance) when the sample and/or the digestion medium contained large amounts of chloride. Among the three candidate reaction gases at their optimized Eonditions, creation of VO+ with oxygen gas delivered the best analyte sensitivity and the lowest detection limit (2.7 ng L-1). Vanadium results obtained from fourteen lake sediment samples and a certified reference material (CRM031-040-1), using two different analytelinterference separation strategies, suggested that the vanadium mono-oxidation offers advantageous performance over the conventional method using NH3 for ultra-trace vanadium determination by ICP-DRC-MS and can be readily employed in relevant environmental chemistry applications that deal with ultra-trace contaminants.Part II: Validation of a modified oxidation approach for the quantification of total arsenic and selenium in complex environmental matrices Spectroscopic interference problems of arsenic and selenium in ICP-MS practices were investigated in detail. Preliminary literature review suggested that oxygen could serve as an effective candidate reaction gas for analysis of the two elements in dynamic reaction cell coupled ICP-MS. An accurate method was developed for the determination of As and Se in complex environmental samples, based on a series of modifications on an oxidation approach for As and Se previously reported. Rhodium was used as internal standard in this study to help minimize non-spectral interferences such as instrumental drift. Using an oxygen gas flow slightly higher than 0.5 mL min-I, arsenic is converted to 75 AS160+ ion in an efficient manner whereas a potentially interfering ion, 91Zr+, is completely removed. Instead of using the most abundant Se isotope, 80Se, selenium was determined by a second most abundant isotope, 78Se, in the form of 78Se160. Upon careful selection of oxygen gas flow rate and optimization ofRPq value, previous isobaric threats caused by Zr and Mo were reduced to background levels whereas another potential atomic isobar, 96Ru+, became completely harmless to the new selenium analyte. The new method underwent a strict validation procedure where the recovery of a suitable certified reference material was examined and the obtained sample data were compared with those produced by a credible external laboratory who analyzed the same set of samples using a standardized HG-ICP-AES method. The validation results were satisfactory. The resultant limits of detection for arsenic and selenium were 5 ng L-1 and 60 ng L-1, respectively.

HG 186.9-187.0 D.E-1

Coarse grained sediment with a few fine grained matrices. Brown sediment with small to medium sized clasts. Clasts range from sub-angular to sub-rounded. Organic material present.

HG 186.9-187.0 D.E.

Coarse grained sediment with a few fine grained matrices. Brown sediment with small to medium sized clasts. Clasts range from sub-angular to sub-rounded. Organic material present. It contains rotation structures throughout as well as edge-to-edge grain crushing. Fine grained clay domains are present and lineations can also be seen. Minor amounts of grain stacking can also be seen.