Seismic Vulnerability of a Historical Masonry Building: A Critical Comparison of Analytical MethodologiesMarco Boscolo Bielo
, 2020, doktorska disertacija
Opis: In these last twenty years a new sensibility for the seismic themes around buildings has emerged from a cultural and technical point of view. As a result of some earthquakes happened in Italy, the promulgation of technical codes has accelerated and, essentially, the standards of Eurocodes were accepted. Consequently, also in the field of historical buildings, a set of specific guidelines have been issued.
In the current state of art, designers are faced with many methods of calculation which the technical codes offer for the assessment of the seismic vulnerability of historical buildings. Designers can use simplified or complex approaches which assume different starting hypothesisto obtain different results.
This thesis aims to investigate the various aspects that the designer meets during the study of the static behaviour of an historical building and the assessment of its seismic vulnerability. A study case is also proposed which regards a Venetian Villa of the XVIth Century located in “Riviera del Brenta” near Venice, with a critical comparison of the results obtained by the application of the various methods of calculation.
The thesis begins with an outline of the historical development of seismic technical codes in Italy and shows how the sensibility has developed in the Italian culture. In particular it is illustrated the difference between the approach of the legislation "a posteriori" (where a “patchy zoning” was generally performed after the occurrence of earthquakes), and the probabilistic approach based on the peaking ground acceleration expected in every Italian site.
The Second and the Third Chapters contain the analysis of the current seismic legislations and in particular the guidelines for historical buildings.
The Chapter 4 provides a general discussion on the masonry and the definition of the mechanical parameters. There are many critical aspects relating to a characterization of models which can realistically represent the historical masonry. They are due to a heterogeneity of variables such as, for instance, the mechanical parameters, the physiological behaviour, the connections, etc.
In the Chapter 5 the basic concepts of seismic behaviour of historical buildings are discussed. In particular the Pagano’s classification of masonry buildings is illustrated with special regard of the following factors: stiffness of horizontal structures; efficiency of “masonry beams”; efficiency of intersection between walls.
From the Chapter 6 the analysis of the study case begins. Here we can find the description of the Venetian Villa (historical evolution of the buildings, description of geometrical characteristics, structural behaviour, etc.
In the Chapter 7, the first level of analysis is performed for the study-case. The method of calculation is contained in the guidelines for historical buildings. A critical discussion of the results is also illustrated.
In the Chapter 8 is performed a linear kinematic analysis based on various possible kinematic mechanisms identified through the study of cracking and the lacks of the structural
The Chapter 9 shows a non-linear static analysis (pushover) for the study-case which refers to two limit behaviour models: the first one with infinitely rigid diaphragms and efficient masonry beams; the second one based on infinitely flexible diaphragms and weak masonry
beams. The results are compared and critically discussed.
In the same way the Chapter 10 and 11 perform a linear static analysis and a linear dynamic analysis for the two models of the Chapter 9, so the results can be compared.
In the last Chapter a general discussion about all the results of the performed analysis is conducted. Here the work points out the conceptual limits of each method of calculation and the possibility of use related to the objectives which must be reached. Finally some design indications related to the results of the analysis are illustrated.
Ključne besede: Seismic, Vulneraility, Analitical, Methodologies
Objavljeno: 02.07.2020; Ogledov: 219; Prenosov: 5
Polno besedilo (128,89 MB)
FUNCTIONALIZED NANOBODIES AS DIAGNOSTIC REAGENTS FOR THE DETECTION OF TOXIC MICROALGAESandra Folarin Oloketuyi
, 2020, doktorska disertacija
Opis: Early detection and monitoring of toxic microalgae remains a challenge that will be necessary to overcome for understanding harmful algal bloom events and their consequent ecological and environmental impacts. Although there are diagnostic techniques based on microscopy, fluorescence in situ hybridization (FISH), DNA microarrays, real-time PCR, sandwich hybridization assays employed for the detection of harmful algal species, such techniques are not suitable for field monitoring and especially for the quantification of Alexandrium minutum due to abundance of cryptic species that cannot be differentiated based on morphology or genomic traits. These approaches are also time-consuming and laborious, hence there is need for alternative simple, rapid and cost-effective methods. In this study, our aim was to develop different nanobody-based capture strategies for the detection of A. minutum. Nanobodies were successfully displayed on the surface of Escherichia coli to facilitate detection of A. minutum. The method is simple and cost effective as no antibody purification step is necessary. As an alternative, A. minutum was quantified by exploiting the peroxidase activity of a G-quadruplex DNAzyme covalently bound to nanobodies and by means of a label free electrochemical immunosensor which exploit nanobodies bound via an irreversible SpyTag-SpyCatcher system as the specific immunocapture element. Such immunosensor was prepared by immobilizing the SpyTagged anti-A. minutum nanobody (SpyTagged C1) by means of a selfassembled monolayer (SAMs) of L- cysteine (L-Cys) displayed onto a gold nanoparticle-modified glassy carbon electrode. Electrochemical Impedance Spectroscopy (EIS) was performed to measure the electrical response of the circuit as the function of A. minutum immunocaptured cells. The results of EIS studies confirmed that the sensing chip detected A. minutum selectively, exhibited a wide dynamic range spanning from 103 to 109 cell L-1 and a limit of detection of 3.1× 103 cell L-1. The immunosensor data can be recorded and then analyzed with a portable potentiostat. Such device can be interfaced with a smart phone, a condition suitable for the rapid A. minutum quantification in situ.
Ključne besede: Alexandrium minutum, nanobodies, bioreagents, immunosensor, toxic microalgae
Objavljeno: 17.06.2020; Ogledov: 325; Prenosov: 14
Polno besedilo (2,95 MB)
DEVELOPMENT OF A NOVEL PCR-BASED ASSAY FOR HIGH-RISK HUMAN PAPILLOMAVIRUS DETECTION AND GENOTYPING IN SELF COLLECTED CERVICOVAGINAL SAMPLES: A NEW POSSIBILITY FOR THE CERVICAL CANCER SCREENINGAlice Avian
, 2020, doktorska disertacija
Opis: Human Papillomavirus (HPV) infection is the causative agent for the invasive cervical cancer and its precancerous lesions, furthermore, there are growing evidences of HPV being a relevant factor in other anogenital cancers as well as head and neck cancers. Most sexually active women become infected with HPV at least once in their lifetime, but less than 10% of women becomes persistently infected, and it is precisely the persistent infection that contributes to the development of cervical cancer.
The preventive effect of cervical cancer screening largely depends in the high women participation and coverage; indeed, a large number of cervical cancers diagnoses normally arise among under-screened and unscreened women. Increase in the screening coverage is essential to improve the effectiveness of cervical screening programmes.
The main purpose of this PhD project was to solve some of the most relevant problems in the cervical cancer screening programmes, as the increase of cost-effectiveness and the amelioration of the screening coverage.
My work was focused on the development and validation of the first Ulisse BioMed S.p.A. product, the HPV Selfy™ test, an innovative PCR-based kit for the direct detection and genotyping of 12 high-risk HPV types (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58 and 59) and 2 possible/probable high-risk (66 and 68), specifically optimized for the analysis of self-collected vaginal specimens.
The core of this innovative test is based on high-resolution melting (HRM) analysis, a recently developed technique for fast, high-throughput post-PCR analysis of variance in nucleic acid sequences, that characterizes the amplicons by studying thermal denaturation of double-stranded DNA. Based on this approach and through the design of different HPV type-specific primer pairs and the development of a specific master mix, unique melting peaks in a single fluorescence channel were obtained, allowing the multiple detection and genotyping of 14 HPV types in a single PCR well.
Three different clinical studies have been carried out to validate the assay on the vaginal self-collected samples with truly amazing results regarding the assay’s performance, but also for self-sampling acceptability by women. Moreover, data collected in these studies suggest a future possible use of this test for the hard-to-reach women, as an alternative of the conventional clinician-collected sample, in order to increase the cervical cancer screening coverage.
Ključne besede: Human Papillomavirus, HPV test, cervical cancer screening, prevention, diagnostic test, High resolution melting, HRM, genotyping, PCR, Self-sampling, clinical validation.
Objavljeno: 17.06.2020; Ogledov: 228; Prenosov: 4
Polno besedilo (37,88 MB)
Managing karst in Coastal British Columbia, CanadaPaul Andre Griffiths
, 2020, doktorska disertacija
Opis: This thesis provides a detailed description and analysis of the system used for managing karst in the forests of coastal British Columbia (BC), where the major land- use activity is industrial forestry. In 2004, BC shifted from a more prescriptive forest management model (the Forest Practices Code) to a less regulated, results-based approach based primarily on the Forest and Range Practices Act (FRPA). The dissertation examines how this results-based management approach for forest resources has worked – or not worked – with respect to karst.
The research focused on the five key realms of an environmental management system as defined by the International Standards Organization (ISO 14001 standard): 1) Legislation and Policy, 2) Planning, 3) Implementation, 4) Checking/Corrective Action, and 5) Management Review. The research also examined the roles of professional reliance (another major foundational element of the FRPA model), karst research, and organizational capacity as external factors which influence on the functioning of the karst management system. A combination of interviews, surveys, document reviews and field observations were used to collect qualitative information relevant to all aspects of the karst management system.
This research reveals inadequacies in all five realms of BC’s current karst management framework, including gaps in legislation, a lack of implementation of existing standards and guidelines, and non-existent effectiveness and compliance monitoring. The results suggest that BC’s shift to ‘self-regulation’ has yielded unsatisfactory results so far for karst resources and has hindered progress toward implementing a fully integrated science-based ecosystem approach to karst management in the study area. Professional reliance failures are identified as one of the key factors contributing to a breakdown of the management system for karst.
This is the first comprehensive study that examines the systems and processes used for managing karst in coastal BC, and consolidates knowledge for government, industry, and others that wish to study or better understand BC’s approach and methods for managing karst. The findings will be useful for private and public forest sector organizations endeavoring to implement fully-functional and effective systems for managing karst in a forestry context. This information may also have more specific applications for managing karst.
Ključne besede: karst ecosystems, protection and management, legislation, policies, standards and guidelines, environmental degradation, forestry impacts, professional reliance, British Columbia, Canada
Objavljeno: 02.03.2020; Ogledov: 443; Prenosov: 17
Polno besedilo (136,27 MB)
Dissecting the role of REEP1 in preventing Tau-mediated neurodegeneration in a D.melanogaster Alzheimer's disease modelAlessio Guglielmi
, 2019, doktorska disertacija
Opis: Tau is natively an unfolded protein that promotes the assembly and the stability of the
axonal microtubules in the central nervous system. Increased formation of Tau protein
aggregates has been causatively implicated in several neurodegenerative diseases called
tauopathies. In the present study, we used the Drosophila melanogaster system to express
the longest isoform of human Tau (2N4R) in the nervous system of adult flies, recreating
the main features of the human pathology. Herein, this Tau-mediated neurodegeneration
model was used as a platform to perform genetic screenings to identify putative modifiers
of Tau toxicity. Our strategy exploited the modulation of genes considered as risk factors of
Alzheimer’s disease (AD), Frontotemporal Dementias and other neurodegenerative
diseases by RNA interference in vivo. This approach allowed us to identify a new gene
which participates in the neuronal response against Tau induced neurotoxicity in
Drosophila: D-Reep1, homologue of human REEP1 gene (h-Reep1). D-Reep1 knockout flies
showed no apparent phenotypes in physiological growing and developmental conditions,
however, they showed peculiar sensitivity to stress conditions. In addition, D-Reep1
knockout enhanced the neurodegeneration mediated by Tau expression in Drosophila eyes.
On the contrary, the overexpression of UAS-D-Reep1 and UAS-h-Reep1 abolished the
typical rough eye phenotype induced by the presence of Tau. The Co-expression of D-Reep1
in Tau backgrounds did not alter the phosphorylation pattern of this protein while, the
presence of D-Reep1 seemed to prevent the formation of Tau aggregates in vivo. Thus, the
data support the idea that D-Reep1 exerts a protective role on Tau induced toxicity which is
independent of its phosphorylation status. In this work, I analysed the mechanisms behind
the neuroprotective role of D-Reep1 and, in particular, I found that REEP1 is involved in the
regulation of the unfolded protein response (UPR) through the PERK-ATF4 cascade within
the ER. By the activation of this pathway, the neurotoxic aggregates of Tau are removed
from Drosophila neuronal tissues rescuing the normal characteristics of the affected
tissues. Evidences also suggest that the activation of autophagy was behind the removal of
Tau aggregates, providing new molecular information about the physiological role of D
Reep1 in the nervous system.
Ključne besede: AD Alzheimer Disease APP Amyloid precursor protein CNS Central Nervous System DM Drosophila melanogaster HSP Hereditary Spastic Paraplegia LN Lewy’s neurite MT Microtubule MAP Microtubule associated protein MT Microtubule/s MTBD Microtubule binding domain NFT Neurofibrillary tangle NP Neuritic plaques PHF Paired helical filament PS1 Presenilin 1 PS2 Presenilin 2 SPG Spastic Paraplegia ThS Thioflavin S
Objavljeno: 06.12.2019; Ogledov: 555; Prenosov: 23
Polno besedilo (2,59 MB)
Photoactive nanocomposite thin films on glass and thermosensitive substratesNives Vodišek
, 2019, doktorska disertacija
Opis: Photocatalysis is a well-known process for the last few decades; it is predominantly used for water and air purification, but also for self-cleaning and antibacterial surfaces. The photocatalytic process is one of the advanced oxidation processes, where semiconductors are mainly used as photocatalysts. The most known and used semiconductor is nanosized titania, which can non-selectively decompose organic matters. One of the side effects of nano TiO2 under UV irradiation is photoinduced hydrophilicity, which furthermore improves the self-cleaning effect. The main drawback of TiO2 is that for its activation UV light is needed, which represents only 5 percent of the solar spectrum. Consequently, actinic irradiance power is low and the response of photocatalyst is limited. Novel approaches are being introduced to improve TiO2 response to visible light such as doping, coupling, modification of surface morphology and others.
The thesis consists of five principal chapters. The first chapter is focused on a short literature review and explanation of some basic terms and principles that are related to this thesis.
The aim of the research was to prepare transparent photocatalytically active thin films on glass and thermosensitive substrates. Four thermosensitive substrates were used: polyvinyl chloride (PVC) foil, polymethyl methacrylate (PMMA) sheet and polyester (PES) fabric coated with a polyvinyl and acrylic coating, with (D1) or without (D2) an additional polyvinylidene fluoride topcoat. The synthesis of films was modified with the introduction of zirconium aiming at improving the photocatalytic activity of the TiO2 films, and the final films had a SiO2 phase present for increased mechanical robustness.
The third chapter is dedicated to the experimental part of the thesis. The detailed process of synthesis is described. The sol-gel process was used to prepare initial Ti-Zr sols, colloidal aqueous solutions with TiO2 and ZrO2 nanoparticles derived from titanium and zirconium alkoxides. The four different Ti-Zr sols were prepared with 0, 5, 10 and 20 molar % of Zr according to Ti. The depositing solution was prepared by mixing Ti-Zr sol, SiO2 sol binder, 1-propanol and 2-propoxyethanol, and then thin films were deposited by dip-coating technique. After the deposition, samples did not require high-temperature calcination since the photocatalytic anatase phase was present already after drying the deposited layers. Coated glass slides were put into a furnace at 150 °C, while samples on thermosensitive substrates were just treated by a heat gun. Samples in powder and thin film form were characterized by UV-Vis, FTIR-ATR and laser beam deflection spectroscopy, X-ray diffraction, scanning electron microscopy and thermal analysis. One of the main aspects of the photocatalyst is its activity. Photocatalytic activity of the thin films was determined, either quantitatively by the formation of fluorescent hydroxyterephthalic acid, one of the first degradation products of terephthalic acid deposit, or qualitatively by a visual-based method where degradation of resazurin ink was observed. Two additional methods were used on glass samples, electron paramagnetic resonance, and degradation of methyl stearate by observing the change in water contact angle under UVA irradiation. Mechanical stability of films on various substrates is an essential factor, describing how successful was immobilization of the photocatalytic materials on the substrate. It was determined by the pencil hardness test, i.e., Wolff-Wilborn method.
In the fourth chapter, titled “Results and Discussion,” the most important part of the thesis is placed. Materials characterization and associated discussion of the results are divided into three major parts: i) powder samples characterization; ii) characterization of thin films on a glass substrate, and iii) characterization of films on thermosensitive substrates.
In the conclusions, the main findings of the Ph.D. research work are summarized. One of our initial hypotheses was disapproved, thin films were not more active with a higher content of zirconium. However, they were more durable. All the obtained films were transparent and photoactive, despite the low-temperature synthesis procedure. Besides commonly used glass substrate, immobilization on thermosensitive substrates was also successful.
Ključne besede: glass substrate, immobilization, nanocomposite, nanomaterials, PES, photocatalysis, PMMA, PVC, self-cleaning surfaces, silica, plastic substrates, thin films, titania, zirconia
Objavljeno: 14.10.2019; Ogledov: 642; Prenosov: 52
Polno besedilo (6,23 MB)
SEARCH FOR NEUTRINOS AT EXTREME ENERGIES WITH THE PIERRE AUGER OBSERVATORYMarta Trini
, doktorska disertacija
Opis: The detection of Ultra-High-Energy (UHE) neutrinos around and above 10 18 eV (1 EeV) can be the key to
answering the long-standing question of the origin of the UHE cosmic rays. The Pierre Auger Observatory
is the largest experiment that can detect the extensive air showers produced when the cosmic rays and
neutrinos interact in the earth’s atmosphere. In particular, with the Infilled array of the Surface Detector
of the Pierre Auger Observatory we can detect sub-EeV neutrino-induced particle showers. In this thesis
we demonstrate that it is possible to discriminate neutrino-induced showers from the background showers
produced by the more numerous nucleonic cosmic rays. The sensitivity to neutrinos is enhanced in the
inclined directions with respect to the vertical to the ground, where cosmic ray-induced showers starting in
the upper layers of the atmosphere are dominated by the muonic component of the shower, while deeply-
penetrating neutrino showers in contrast exhibit a large electromagnetic component. Based on this idea in
this thesis we have developed a search procedure for UHE neutrinos that consists on selecting inclined
events in the Infilled array of the Pierre Auger Observatory in which the signals in the water-Cherenkov
stations are spread in time, characteristic of the presence of electromagnetic component in the shower. We
have established a complete chain of criteria to first select the inclined events among the sample of all
events triggering the Infilled array, and then identifying those that have a large electromagnetic component
at ground, and hence can be considered as neutrino candidates. We have identified a single variable, the
so-called area-over-peak averaged over all of the stations in each event, as a suitable observable for neutrino
identification purposes. The neutrino selection was established using extensive Monte Carlo simulations of
the neutrino-induced showers in the Infilled array of Auger as well as a fraction of the data assumed to
be totally constituted of background nucleonic cosmic rays. Using these neutrino simulations we have also
computed the exposure of the Infilled array to UHE neutrinos in the period 1 January 04 - 31 December 2017.
Associated systematic uncertainties on the exposure are also described. Expecting no candidate neutrinos in
the period up to 31 December 2017, and adopting a differential neutrino diffuse flux dN ν /dE ν = k E ν −2 in
the energy range from 0.05 to 1 EeV, we have obtained a 90% C.L. upper limit on the all neutrino flavor,
k 90 < 7.97 × 10 −8 GeV cm −2 s −1 sr −1 .
Ključne besede: astroparticles, astrophysical neutrinos, cosmic rays showers, Pierre Auger Observatory, Infilled
Objavljeno: 03.10.2019; Ogledov: 1231; Prenosov: 31
Polno besedilo (6,02 MB)
Factors influencing collected quantities of municipal solid wasteSlavica Schuster
, 2019, doktorska disertacija
Opis: The first part of the survey, using the survey (enclosed with the annexes), tried in various ways to identify samples and facts by which we could predict the amount of waste collected according to some social and economic indicators of all municipalities of the Republic of Slovenia. Data were collected through surveys to determine the way we work and think about the collection of waste and landfills. For my research I have selected the Republic of Slovenia and its 212 municipalities. It is precisely Slovenia that in many respects represents a bright spot in waste management and management.
In the second part of the research we used statistical methods to find the factors that influence the increase of waste. From the results we have proved our hypotheses and one of the most important is how the average age of people influences the increase of waste. We did the research over a period of 6 years (2012-2017). Where, by comparison, we find that they do not change significantly in structure by age, but of course socio-economic indicators such as average age change.
In the last section, we compared the growth of waste collected with GDP growth. We show that with the growth of GDP, the amount of waste collected also increases. The research is also based on the Republic of Slovenia and beyond. In the second part of the last survey we also included data for EU Member States (28 of them).
Ključne besede: waste, waste management, amount of waste, socio-economic impacts, Slovenia, wild-landfills, GDP
Objavljeno: 02.10.2019; Ogledov: 682; Prenosov: 31
Polno besedilo (3,88 MB)
HOLISTIC GEOMORPHOLOGICAL SPATIAL ANALYSIS OF KARST ON KRK ISLANDEla Šegina
, 2019, doktorska disertacija
Opis: The intriguing spatial variability of surface features on Krk Island has stimulated the research of this karst area located in the coastal zone of the Dinaric karst in Croatia. Field inspection, ortho-rectified aerial photos (0.5m resolution) and a topographic map (1:5,000) were used for the detection and delineation of detectable karst surface features appearing on the island with the area of 405.5 km². This method resulted in the identification of several yet undefined types of surface features occurring on karst, requiring the revision of the existing classification and re-establishment of a new classification system compatible with the particular field reality. Several morphologic and distributive parameters that had been calculated for each re-classified type of surface feature provided insight into the surface features elementary characteristics, their spatial variability and the correlation to the other types of surface features and to the recent karst relief. This analysis based on a large, accurate dataset, contributed to the general knowledge on karstic surface features, the conditions of surface features in Dinaric karst and to the understanding of the karst surface evolution on Krk Island.
Ključne besede: karst geomorphology, GIS, spatial analysis, Krk Island, Dinaric karst, Adriatic Sea
Objavljeno: 13.09.2019; Ogledov: 939; Prenosov: 63
Polno besedilo (17,12 MB)
DETEKCIJA KOVINSKIH KOMPLEKSOV IN ORGANOKOVINSKIH SPOJIN V VZORCIH IZ OKOLJA S SPEKTROMETRIJO TERMIČNIH LEČLeja Goljat
, 2019, doktorska disertacija
Opis: Environmental pollution is one of the greatest challenges that the world is facing today. Toxic compounds, such as pesticides, allergens, pharmaceuticals, toxins and heavy metals are widely present in the air, water and soil, and can affect the health of people and animals even in small quantities, as well as they may cause long- or short-term damage in plants [Hill, 1997].
Heavy metals (mercury, arsenic, cadmium…) are widely spread in the environment. They derive from a number of sources, including mining, industrial wastes and vehicle emissions [Tchounwou et al., 2012]. They are easily incorporated into biological molecules and exert their toxic effects by displacing essential metals of a lower binding power in biologically active molecules or by acting as noncompetitive inhibitors of enzymes, affecting neurological, reproductive, renal and hematological systems [Sunil D’Souza et al., 2003; Heavy-Metal Pollution, 2018]. Metals form countless compounds (e.g. metal complexes and organometallic compounds) which are essential for living organisms (vitamin B12, hemoglobin, chlorophyll) and/or have a wide range of applications in industry and other areas, including analytical chemistry. Because of the potential risk which toxic metals represent to the living organisms and also because of the importance of some essential metals, different analytical techniques and detection methods have been developed for studies of their occurrence, fate and concentration in the environment and in organisms. However, providing a required sensitivity for determination and speciation of different metals and their compounds, especially in small- volume samples is still a challenge.
Therefore, general objectives of this dissertation were development of novel analytical methods for sensitive, reliable and fast determination of metal species, based on highly sensitive optothermal technique thermal lens spectrometry (TLS), which can be used as detection tool following colorimetric reaction of a selected metal ion or for direct detection of colored organometallic compounds.
This dissertation is composed of the following chapters: introduction, research goals, theoretical background, results and discussion, conclusion and references. The core of this dissertation is presented in the fifth chapter (results and discussion), which is divided into three parts. They separately cover development of methods for determination of iron redox species, pyoverdine and Fe-pyoverdine complexes and mercury. Pyoverdine is a siderophore, excreted by a certain bacteria in order to scavenge iron in the environment and is closely related to the chemistry of iron in such biological systems. Therefore, the first two parts are closely related.
Procedures for batch mode thermal lens microscopy (TLM), flow-injection thermal lens sprectrometry (FIA-TLS) and µFIA-TLM (flow injection and TLS detection in microspace) were developed for Fe(II) and Fe(III) determination, based on colorimetric reaction of Fe(II) with 1,10-Phenanthroline. All these procedures were focused on cloudwater examination with a tendency to minimize sample consumption but at the same time preserve low limits of detection (LOD) and limits of quantification (LOQ). TLM measurements with highly collimated probe beam were performed in a 100 μm optical path length cell (40 µL volume), which resulted in a considerably smaller sample volume requirement (500 µL in total) and consumption, as compared to UV-Vis spectrophotometry, which required at least 25 mL of sample due to large volume (almost 30 mL) of the 10 cm optical path-length sample cell. LODs for mode-mismatched TLM were 0.16 and 0.14 µM for Fe(II) and Fe(total) (sum of Fe(II) and Fe(III) concentrations), respectively, while LODs for UV-Vis spectrophotometry were 0.01 µM for both Fe(II) and Fe(total). By using the mode mismatched TLM we were able to detect concentrations corresponding to absorbances as low as 1.5 × 10-5, while the lowest absorbance detectable on the UV-Vis spectrophotometer corresponded to 1.1 × 10-3, despite the use of the 10 cm optical path-length cell.
Another important step in the development of new methods for Fe(II) and Fe(III) determination was the use of TLS detection in FIA (FIA-TLS). By injecting 50 µL of the sample into the FIA-TLS system, cca. 10 times lower LODs were achieved (1 × 10-3 µM for Fe(II) and 8 × 10-4 µM for Fe(total)), as compared to the UV- Vis spectrophotometry.
Nevertheless, the development of μFIA-TLM method, with on-line colorimetric reaction for Fe(II) and Fe(III) determination is considered as the most important achievement of this study. The results show that despite 100 times shorter optical path length and low sample consumption (3 µL of each sample/injection) compared to UV-Vis spectrophotometry, LODs for µFIA-TLM were 0.10 and 0.07 μM for Fe(II) and Fe(total) respectively, which is sufficiently for cloudwater analysis, since concentrations, lower than 0.1 μM are not expected [Parazols et al., 2006; Deguillaume et al., 2014]. Linear range for Fe(II) and Fe(III) determination by μFIA-TLM was between 0.1 and 70 µM. To test the accuracy of this method, artificial cloudwater was prepared, spiked with different amounts of Fe(II) and Fe(III) and analyzed for iron content by µFIA-TLM and UV-Vis spectrophotometry. Good agreement was observed between the two methods. To ascertain the ruggedness of the method 7 (or more) replicate determinations at two different concentrations for both, Fe(II) and Fe(total) in artificial cloudwater were carried out on day 1 (replicates were measured instantly after fortification), day 2 and day 5. A student’s t-test (p=0.05) was applied to compare 3 sets of obtained data (day 1, day 2 and day 5) and showed that sets are not significantly different from each other. Considering very low sample volume requirement of µFIA-TLM, this should be the method of choice for determination of Fe(II) and Fe(III) in investigations of processes in cloudwater, where multiparameter analysis is desired (determination of other ions, ligands, microbial counts, etc.). When larger sample volumes are available, FIA-TLS can be used for accurate determination of iron species at lowest concentration levels.
High performance liquid chromatography (HPLC) was applied for separation and detection of pyoverdine (PVD), produced by Pseudomonas fluorescens 36b5, a bacterial strain isolated from the aqueous phase of clouds at the Puy de Dôme station (1465 m, France). Reversed-phase (RP) chromatography (RP-18 chromatographic column Hypersil gold), hydrophilic interaction liquid chromatography (HILIC) (ZIC®-Hilic column) and three different detection systems
(diode-array (DAD), spectrofluorimetry (FLD) and TLS) were tested for their performance in separation and determination of pyoverdines and corresponding complexes of pyoverdine with iron (Fe(III)-PVDs).
PVDs and Fe(III)-PVD complexes could not be separated and quantified by applying HILIC technique, therefore it was concluded, that HILIC is not suitable for HPLC-DAD and also not for HPLC-TLS, since the method should offer a simultaneous sensitive detection of free PVDs as well as Fe(III)-PVD complexes in a single chromatographic run.
Since pyoverdine standards were available only as a mixture of several different forms of PVDs, whereby the exact composition was unknown, the quantification of each of the four major specie (two fluorescent PVDs and two nonfluorescent Fe(III)-PVDs) in the standard, which was obtained from Université Clermont Auvergne, Institut de Chimie de Clermont-Ferrand, was performed. When applying Hypersil gold column, a linear correlation between fluorescence intensity and absorbance of each component was observed in a concentration range 3–24 µg/mL, whereby LODs were estimated to be 0.03–0.04 µg/mL for each of the major PVD species (HPLC-DAD). Even though HPLC-FLD method provided cca. 100 times lower LODs, it is not the method of choice for determination of PVD species in cloudwater, because it does not allow detection of PVD complexes with Fe(III). When comparing HPLC-TLS and
HPLC-DAD, LODs were 5 to 8 times lower in case of HPLC-TLS, which was a significant improvement. Furthermore, recoveries (89–111 %) at two concentration levels of four PVD species in two independent samples, showed good reliability of the method.
Almost all mercury in uncontaminated drinking-water is thought to be in the form of Hg2+ [WHO, 2010]. Therefore, the method for Hg2+determination based on colorimetric reaction with triamterene, described originally by Al-Kady and Abdelmonem was further investigated in this study, as well as the possibilities of application of this reaction for Hg2+ determination by TLS. The stoichiometry of the complex formation was determined by the method of continuous variations and saturation experiment, suggesting formation of the complex with the formula Hg2-triamterene. The obtained value of the molar absorption coefficient was 9988 Lmol-1cm-1 at 403 nm, which significantly contradicts the existing data in literature, which reports the molar absorption coefficient of 5.32 × 104 Lmol-1cm-1 [Al-Kady and Abdelmonem, 2013]. Even though the spectrophotometric results were not encouraging for triamterene as colorimetric reagent for Hg2+ determination, it was further investigated for its performance in TLS system. Fe(II)-1,10-phenanthroline (ferroin) was used for comparison, because it was well studied for TLS applications previously. The results showed that Hg2-triamterene in solutions was degraded when it was exposed to the light of the excitation beam. Due to the lower molar absorptivity than reported in literature, fotodegradation and unfavorable complex stoichiometry, triamterene was not confirmed as a suitable colorimetric reagent for highly sensitive Hg2+ determination by TLS.
In summary, this dissertation investigates alternative approaches for analysis of metal complexes and organometallic compounds in small-volume environmental water samples. Methods, which were developed in this research, could potentially serve as improvements of existing technologies, to facilitate analysis of such samples, by offering simple handling of samples and superior sensitivity over the UV-Vis spectrophotometry.
Ključne besede: thermal lens spectrometry, thermal lens microscopy, high performance liquid chromatography, microfluidics, metal complexes, organometallic compounds, iron, pyoverdine, mercury
Objavljeno: 05.09.2019; Ogledov: 675; Prenosov: 40
Polno besedilo (3,65 MB)