Repozitorij Univerze v Novi Gorici

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Avtorji:ID Prosenc, Franja (Avtor)
ID Franko, Mladen (Mentor) Več o mentorju... Novo okno
Datoteke:.pdf Disertacija_april2017.pdf (27,08 MB)
MD5: C1BEDD6FB2EC7F722696F6654929B3D8
Jezik:Angleški jezik
Vrsta gradiva:Doktorsko delo/naloga
Tipologija:2.08 - Doktorska disertacija
Organizacija:FPŠ - Fakulteta za podiplomski študij
Opis:Water security and quality are a global issue of concern, which have recently become alarming due to the growth of the human population, industrialisation and expanded agricultural activities. Biologically active compounds, such as pharmaceuticals and personal care products can have major adverse effects on aquatic organisms, and are therefore one of the biggest threats in water quality. Another major concern is the spread of waterborne pathogens, including multidrug resistant (MDR) bacteria, which can cause serious illnesses in humans. In order to maintain water abundance and quality, it is necessary that adequate wastewater treatment and analytical techniques allowing for sensitive and fast-response detection of water hazards are in place. Conventional (waste)water treatment technologies often fail to adequately remove all of the water hazards detailed above. Moreover, conventional analytical techniques currently used in water quality control are, although highly selective and sensitive, time-extensive, with throughput of merely 2 to 3 samples per hour, excluding the time for sample preparation. With respect to these drawbacks, research was proposed to explore new approaches for degradation of recalcitrant compounds, inactivation of microorganisms, and fast screening methods, which are listed in the second chapter of this dissertation as research objectives. In the third chapter, an extensive theoretical background on the hazards found in aquatic environment, namely pharmaceuticals and waterborne pathogens, is given. Pharmaceuticals enter the environment through several routes (disposal of unused medication via the toilet, pharmaceuticals passing through the human body unchanged/slightly transformed, animal excretions of pharmaceutically active compounds, insufficient wastewater treatment, etc.); therefore, traces of pharmaceuticals have repeatedly been reported in surface waters, groundwater, wastewater effluents, and even drinking water. Iodinated contrast agents (ICAs), as the compounds of interest in this project are further described. ICAs are eliminated from the human body practically unchanged; therefore a large proportion of them end up in municipal and hospital wastewater, where they can be present in concentrations of up to 2.4 g/L. Their ecotoxicity, degradation attempts, as well as detection monitoring in the environment are reviewed within the chapter. Additionally, waterborne pathogens, which account for 2.2 million deaths per year, are reviewed in this chapter, with emphasis on multidrug-resistant (MDR) bacteria. Although MDR infections are mostly prevalent in hospital environments, the presence of MDR bacteria in the environment is not a rarity. A high percentage of bacterial isolates in waters have been shown to be of an MDR phenotype. The theoretical background in analytical methods in water quality monitoring is also given in this chapter. Vanguard and rearguard techniques are explained, the first offering simple, cheap, and rapid sample screening, but sacrificing sensitivity and selectivity, whereas the second providing the highest quality information, excellent sensitivity and selectivity, but in expense of complicated and timely sample handling and high-cost instruments. By combining the two techniques the benefits of both can be exploited in a single system. The basic principles of thermal lens spectrometry (TLS) and its miniaturised version - the thermal lens microscopy (TLM) as fast screening methods providing high sensitivity are further explained, and their practical applications are reviewed. Furthermore, composite materials have recently been finding applications in water treatment technologies, as filter materials, adsorptives for pollutants, catalysts for degradation reactions, and disinfectants. The applications of three main types of composites: synthetic composites, biocomposites, and nanocomposites, are also reviewed within this chapter. The core of this dissertation is presented in the fourth and the fifth chapter, which examine two separate approaches for water treatment, as well as analytical methods for fast screening purposes. The fourth chapter is investigating options for degradation of iodinated X-ray agents (ICAs), namely diatrizoate, through biodegradation with extracellular enzymes of white rot fungus Dichomitus squalens, and chemical oxidation with manganese(III) acetate. Enzymatic degradation with laccase (Lac) and manganese peroxidase (MnP) at low enzymatic activities was unsuccessful, whereas at approximately 3-times higher activities the enzymes were capable of 60 % degradation in 12 days. Chemical oxidation of diatrizoate with manganese(III) acetate resulted in 85 % degradation in 12 days. Moreover, the suitability of microfluidic flow injection analysis coupled with thermal lens microscopy (μFIA-TLM) as a fast screening method for diatrizoate degradation was examined. The degradation was monitored through the release of iodide from the diatrizoate molecule. μFIA-TLM proved to be a preferable method over UV-Vis spectrophotometry, due to its higher sensitivity, sample throughput, and simple sample handling. Limit of detection (LOD) for μFIA-TLM method was estimated to be 0.14 µM in a 100 µm channel, which is 9 times lower than LOD obtained in UV-Vis measurements. In addition to μFIA-TLM and UV-VIS, high-pressure liquid chromatography (HPLC) was used to monitor the remaining parent compound in the reaction mix. In the fifth chapter, the second water treatment approach is described. This includes synthesis of biocomposite materials from cellulose (CEL) and keratin (KER), with metal (Ag0, AgCl, Au0) nanoparticles (NPs). Materials were characterised for presence, species, and size of NPs with X-ray diffraction (XRD) and with scanning electron microscopy (SEM). Nanoparticles were confirmed to be of expected species, with sizes as follows: 6.3 ± 0.5 nm for Au NPs, 12 ± 2 nm for Ag NPs, and 22 ± 1 nm for AgCl NPs. In order to evaluate antibacterial properties of the materials, contact tests with gram-negative (Escherichia coli and Pseudomonas aeruginosa) and gram-positive bacteria (Staphylococcus aureus, Methicillin-resistant S. aureus (MRSA), and Vancomycin-resistant Enterococcus faecalis (VRE)) were conducted. Direct-contact assay over 24 hours showed a dose and species-dependent antibacterial activity of [CEL:KER + Ag NPs] materials. The highest potency against the selected bacteria (up to 6-log of reduction) was observed for the material with 500 mg of Ag NPs. AgCl NPs appeared to be less potent than Ag NPs, whereas Au NPs exhibited antibacterial activity only against MRSA and VRE. In addition, antiviral properties of materials were investigated on selected bacteriophages (MS2, phiX174, and fr). However, biocomposite materials with 500 mg of Ag NPs and AgCl NPs, as well as, 240 mg of Au NPs did not exhibit any activity against selected bacteriophages. Biocompatibility with human fibroblasts was evaluated through a direct contact assay for 3 and 7 days of exposure. High concentrations of metal NPs turned out to be cytotoxic for human fibroblasts, whereas the amount of 69 mg of Ag NPs in [CEL:KER] was low enough not to affect the viability of the fibroblasts after 3 days of exposure. Composites with Ag NPs and AgCl NPs were also tested for leachability of NPs out of the materials. Both types of NPs were leaching out in two different forms, as silver ions, and as colloidal silver. Leaching of ionic silver from both materials stabilised after 3 days, whereas colloidal silver was still leaching out on the 7th day. The overall percentage of the total silver (ionic + colloidal) leached was only 0.04 % of silver incorporated in the material. In summary, this dissertation investigates alternative approaches for water treatment technologies, which could potentially serve as unit improvements of existing technologies, or as on-point pre-treatment technologies to facilitate further conventional water treatment techniques. It also demonstrates the suitability of μFIA-TLM for fast screening measurements in aquatic samples, offering high sample throughput, simple handling of the samples and superior sensitivity over the UV-Vis spectrophotometry.
Ključne besede:Antibacterial biocomposites, nanomaterials, water treatment technologies, multidrug-resistant bacteria, iodinated contrast agents, thermal lens microscopy, flow-injection analysis, microfluidics
Kraj izida:Nova Gorica
Založnik:Univerza v Novi Gorici
Leto izida:2017
Št. strani:159
PID:20.500.12556/RUNG-3051-6f3cb0a1-df01-3090-f835-8cec6cc4dabc Novo okno
COBISS.SI-ID:4774907 Novo okno
Datum objave v RUNG:26.04.2017
Število ogledov:5851
Število prenosov:207
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Jezik:Slovenski jezik
Opis:Kakovost in zadostna količina vode sta v zadnjem času zaradi naraščajočega prebivalstva, industrializacije in vse bolj razširjenega kmetijstva, zaskrbljujoči na globalni ravni. Za zagotavljanje teh ciljev je potrebno zagotoviti zadovoljivo stopnjo tretiranja odpadnih voda ter razviti analitske tehnike, ki zagotavljajo občutljivo in hitro detekcijo nevarnih agensov v vodah. Eno večjih nevarnosti predstavljajo farmacevtske učinkovine in izdelki za osebno nego, saj vplivajo na kakovost vode, ker so pogosto biološko aktivni in lahko povzročijo neželene učinke na vodnih organizmih. Drug velik problem kakovosti vode so patogeni organizmi, ki se prenašajo z vodo, vključno z antibiotsko rezistentnimi bakterijami ter njihovim potencialnim širjenjem v okolje. Konvencionalne tehnologije tretiranja (odpanih) voda pogosto nezadostno odstranijo zgoraj izpostavljene nevarne snovi in organizme. Tudi analitske tehnike, ki se običajno uporabljajo pri nadzoru kakovosti vode so kljub selektivnosti in občutljivosti, zamudne, z zmožnostjo analize le 2 do 3 vzorcev na uro (izključujoč čas za pripravo vzorcev). Zaradi zgoraj navedenih pomanjkljivosti so v tej disertaciji raziskani novi pristopi k razgradnji odpornih spojin, inaktivaciji mikroorganizmov ter razvoju novih in hitrih presejalnih analitskih metod. Ti so v drugem poglavju disertacije navedeni kot raziskovalni cilji. Za boljše razumevanje problematike so v tretjem poglavju podane obsežne teoretične osnove o nevarnih agensih, ki se pojavljajo v vodnem okolju, s povdarkom na farmacevtskih učinkovinah ter patogenih organizmih, ki se prenašajo z vodo. Farmacevtske učinkovine vstopijo v okolje po različnih poteh (odstranjevanje neuporabljenih zdravil v stranišče, zdravila, ki zapustijo človeško telo nespremenjena ali le malce transformirana, živalski iztrebki, ki vsebujejo veterinarska zdravila, nezadostno tretirana odpadna voda, ipd.), zato so bile farmacevtske učinkovine v sledovih najdene v površinskih in podzemnih vodah, odpadnih vodah ter občasno tudi v pitni vodi. Nadalje so v tem poglavju opisana jodirana kontrastna sredstva, ki smo jih v tem projektu preučevali kot modelne spojine. Jodirana kontrastna sredstva se izločijo iz človeškega telesa praktično nespremenjena, zato velik delež teh konča v komunalnih in bolnišničnih odpadnih vodah, kjer so lahko prisotna v koncentracijah do 2.4 g/L. V istem poglavju je opisana tudi ekotoksičnost teh spojin, poskusi njihove razgradnje, ter njihova detekcija in monitoring v okolju. V poglavju so obravnavani tudi patogeni organizmi, ki se prenašajo z vodo ter so odgovorni za povzročitev 2,2 milijona smrtnih primerov na leto, s povdarkom na antibiotsko rezistentnih bakterijah. Čeprav so okužbe z antibiotsko rezistentnimi bakterijami primarno problem v bolnišnicah, prisotnost teh bakterij v naravnem okolju ni izjema, saj je velik odstotek bakterijskih izolatov iz naravnih voda kazal fenotip rezistence na več antibiotikov hkrati. Še ena obravnavana tema v tem poglavju so teoretične osnove analitskih metod, uporabljenih pri monitoringu kakovosti vode. Te zajemajo “vanguard” in “rearguard” tehnike. Prve ponujajo preposto, poceni in hitro presejanje vzorcev, vendar na račun nižje selektivnosti in občutljivosti, medtem ko slednje zagotavljajo rezultate najvišje kakovosti, odlično občutljivost in selektivnost, vendar je rokovanje z njimi zapleteno, priprava vzorcev je zamudna in potrebni so dragi inštrumenti. Z združitvijo obeh vrst tehnik je v enem sistemu mogoče združiti prednosti obeh. Opisani so tudi osnovni principi spektrometrije s toplotnimi lečami (TLS) ter njene miniaturne verzije – mikroskopije s toplotnimi lečami (TLM), kot primer hitre presejalne metode z visoko občutljivostjo. Zajet je tudi pregled objav s praktičnimi aplikacijami le teh. Nazadnje pa so v poglavju obravnavani tudi kompozitni materiali, ki se v zadnjem času uporabljajo kot alternativne metode pri tretiranju (odpadnih) voda, in sicer kot materiali za filtre, adsorbenti za onesnažila, katalizatorji pri reakcijah razgradnje ter kot sredstva za dezinfekcijo. V tem poglavju je podan predgled aplikacij treh glavnih tipov kompozitnih materialov: sintetičnih kompozitov, biokompozitov in nanokompozitov. Osrednji del te disertacije je predstavljen v četrtem in petem poglavju, ki vključujeta dva ločena pristopa tretiranja voda, kot tudi analitske metode za hitro in občutljivo zaznavo spojin. V četrtem poglaviju so raziskane možnosti za razgradnjo jodiranih kontrastnih sredstev, natančneje diatrizoata, z biorazgradnjo z izvenceličnimi encimi glive bele trohnobe Dichomitus squalens, ter s kemijsko oksidacijo z manganovim(III) acetatom. Encimska razgradnja z lakazami in mangan peroksidazami je bila pri nizkih encimskih aktivnostih neuspešna, medtem ko so encimi s 3-krat višjimi aktivnostmi bili sposobni 60 % razgradnje v 12 dneh. S kemijsko oksidacijo diatrizoata z manganovim(III) acetatom pa smo dosegli 85 % razgradnjo v 12 dneh. Preverili smo tudi ustreznost mikrofluidne injekcijske analize z mikroskopijo s termičnimi lečami (μFIA-TLM) za hitro zaznavo razgradnje diatrizoata. Razgradnjo smo preverjali preko jodida, odcepljenega iz molekule diatrizoata. Ugotovili smo, da je μFIA-TLM metoda primernejša kot UV-Vis spektrofotometrija, saj je občutljivejša in omogoča analizo večih vzorcev v danem času ter zagotavlja preprostejše rokovanje z vzorci. Za μFIA-TLM metodo smo ocenili mejo detekcije (LOD) 0,14 µM v 100 µm mikrokanalu, kar je 9× nižje kot ocenjena LOD za spektrofotometrične metode. Poleg μFIA-TLM in spektrofotometrije smo razgradnjo spremljali tudi z visokotlačno tekočinsko kromatografijo (HPLC), kjer smo spremljali preostanek diatrizoata v reakcijski mešanici. V petem poglavju je predstavljen drugi pristop k tretiranju voda: sinteza biokompozitnih materialov iz celuloze (CEL), keratina (KER) in kovinskimi nanodelci (Ag0, AgCl, Au0). Material je bil okarakteriziran z rentgensko difrakcijo (XRD) ter z vrstično elektronsko mikroskopijo (SEM), s katerima smo preverjali prisotnost, zvrst in velikost nanodelcev. Potrdili smo, da so bili nanodelci pričakovanih zvrsti ter da so bile njihove velikosti sledeče: 6,3 ± 0,5 nm za Au NPs; 12 ± 2 nm za Ag NPs in 22 ± 1 nm za AgCl NPs. Protibakterijske lastnosti materialov smo ocenili s kontaktnim testom na gram-negativnih (Escherichia coli in Pseudomonas aeruginosa) ter gram-pozitivnih (Staphyloccocus aureus, proti meticilinu odporni S. aureus (MRSA) in proti vankomicinu odporni Enterococcus faecalis (VRE)) bakterijah. 24-urni kontaktni test je pokazal, da so protibakterijske lastnosti [CEL:KER + Ag NPs] materialov odvisne od koncentracije in zvrsti nanodelcev. Najvišjo aktivnost (do 6-stopenj logaritemskega zmanjšanja) smo izmerili pri materialu z 500 mg Ag0 nanodelcev. Nanodelci srebrovega klorida (AgCl) so bili manj aktivni kot nanodelci elementarnega srebra (Ag0), medtem ko so bili zlati (Au) nanodelci aktivni le proti MRSA in VRE bakterijam. Na izbranih bakteriofagih (MS2, phiX174, in fr) smo raziskali tudi protivirusne lastnosti materialov, vendar pa biokompozitni materiali z 500 mg Ag in AgCl, kot tudi 240 mg Au nanodelcev, niso kazali protivirusnih lastnosti proti izbranim bakteriofagom. Biokompatibilnost s človeškimi fibroblasti smo ocenili s kontaktnim testom, kjer smo materiale izpostavili kulturi fibroblastov za 3 in 7 dni. Visoke koncentracije nanodelcev so bile citotoksične za človeške fibroblaste, medtem ko so bili fibroblasti, po 3 dneh inkubacije z materialom z 69 mg Ag NPs, viabilni in primerljivi s kontrolo. Pri kompozitih s srebrovimi (Ag0 in AgCl) nanodelci smo testirali izluževanje nanodelcev iz materialov. Iz obeh materialov so se izluževali nanodelci v dveh oblikah, kot srebrovi ioni ter kot koloidno srebro. Izluževanje ionskega srebra se je pri obeh materialih stabiliziralo po 3 dneh, medtem ko se je koloidno srebro še vedno izluževalo na 7. dan. Skupno izluženo srebro (ionsko + koloidno) je predstavljalo zgolj 0.04 % srebra vključenega v material. V celoti disertacija razišče nekatere alternativne metode tretiranja voda, ki bi lahko potencialno služile izboljšavi posameznih enot obstoječih tehnologij, ali pa tretiranju na izvoru onesnaženja, kar bi olajšajo nadaljnje konvencionalno tretiranje. Prav tako v disertaciji dokažemo ustreznost μFIA-TLM tehnike kot hitre presejalne metode za vodne vzorce. Pokazali smo, da je μFIA-TLM visoko-učinkovita metoda, ki omogoča preprosto rokovanje z vzorci in zagotavlja izredno visoko občutljivost v primerjavi z spektrofotometrijo.
Ključne besede:Protibakterijski biokompoziti, nanomateriali, tehnologije čiščenja voda, multirezistentne bakterije, jodirana kontrastna sredstva, mikroskopija s termičnimi lečami, pretočna injekcijska analiza, mikrofluidika