The detection and study of biologically active compounds in environmental processes and samplesMojca Žorž Furlan
, doctoral dissertation
Abstract: Environmental pollution in the 21th century still represents a global problem for human and animal health. Despite general awareness about released substances and their degradation products their fate and possibilities of removal are not well investigated. Even though the chemicals released are dispersed and diluted in water cycles, their poor biodegradability and/or strong accumulation can result in the intoxication of exposed organisms. Similarly, as a part of the environment, food can get contaminated by bioactive substances during different steps of preparation. Not only artificial compounds such as pesticides or pharmaceuticals, but also natural toxins enter the food chain and impact negatively on humans' and animals' health. In addition, the activity of some bacteria can influence the production of amines from amino acids after fermentation, to which the human body responds with several symptoms of intoxication.
Several analytical methods for the determination of trace levels of broad range contaminants have been developed. Due to the largely robust, selective and sensitive features of the conventional (rearguard) techniques, they represent the first choice for analysing multiple organic compounds in frequently very complex matrices. However, screening (vanguard) methods are paving the way in the chemical analytics as a solution that provides simplicity and rapid analytical responses with binary (yes/no) answers. They require little or no sample treatment as well as more economically-efficient instrumentation. The combination of vanguard-rearguard analytical strategies hence offers a compromise between classical analytical figures of merit and productivity-related characteristics.
In the first part of our research feasibility studies for the application of TLS and/or TLM in novel analytical methods for the determination of lipid-lowering drug atorvastatin and a mycotoxin ochratoxin A . The survey on atorvastatin performed spectrophotometrically has shown a decrease of ATV-sulpho-vanillin product at the wavelength of its maximum absorbance after dilution by organic solvent, which was investigated due to the possible increasing of the method sensitivity. As the predicted LODs that could be obtained by TLM (0.3 mg/L) could not reach the concentration of ATV usually present in the environment (ng/L-g/L) further experiments on this subject were therefore not justified. On the other hand, the ELISA assay for the determination of ochratoxin A was performed. In case of μFIA-TLM, the measurements were influenced by high background signal resulting in high LODs of TLM (470 pg/mL), which is known as a background limited technique. It was estimated that the LODs of standard ELISA assay could not be significantly improved, therefore no further research was conducted in this direction.
In the second part of the dissertation, a sensitive rearguard system by coupling HPLC and TLS for the determination of biogenic amines in wine samples was developed. Putrescine, cadaverine, histamine and tyramine were separated and detected on a HPLC-TLS system after derivatization by dabsyl chloride. The method was optimized in terms of chromatographic conditions and in terms of TLS parameters. Also, the sensitivity of the newly developed method was evaluated by comparing the TLS detection with DAD detection in terms of LOD values, where TLS showed 3.6-fold improvement compared to DAD. Afterwards, the standard addition calibration was performed and evaluated for its recoveries (86−117%) in the determination of the four BAs. The applicability of the novel method was tested by the analysis of real white and red wine samples and by comparing the results to the standard HPLC-FL method and concentrations of BAs in wine samples were in good accordance. In addition, the dabsylated BAs showed better stability compared to the OPA derivatives as they have not lost the peak intensity after 17h of storage.
In the third part, a vanguard system for detection of the overall biogenic amines concentration was developed by employing μFIA-TLM. Initially, NH4Cl standard solutions were applied in the indophenol reaction for batch mode, off-line μFIA-TLM and in an on-line indophenol formation for μFIA-TLM detection. By adding 50 % of EtOH to indophenol we obtained 9-fold improvement. In addition, indophenol showed good stability under TLM conditions. We optimized the microfluidic and TLM parameters in the off-line and on-line indophenol reaction. The addition of 5% ethanol to the reagent in the on-line reaction resulted in the 3-fold improvement of the signal-to-noise ratio. Further on, the overall reaction, including the enzymatic and the following indophenol reaction, was optimized by choosing the optimal buffer (pH=7, 0.5 M) and alkaline conditions (2M NaOH). The influence of interferences from amino compounds was also evaluated and discussed. The off-line and on-line μFIA-TLM were evaluated by their performance characteristics. The LOD for ammonia detection reached 2.3 μM and the applicability in ammonia detection in water samples was discussed. Similar LOD of 3.2 μM was obtained for the overall concentration of BAs and LOD of 3.8 μM for histamine, which is more than 4-folds lower value as the lowest suggested limits of intake for histamine in wine samples (2 mg/L; 18 μM). Finally, an immobilization procedure on magnetic nanoparticles was developed for the possible implementation of the selected enzyme in a miniaturized biosensor.
Keywords: thermal lens spectrometry, thermal lens microscopy, high performance liquid chromatography, microfluidics, biogenic amines, microbial transglutaminase, indophenol (Berthelot) reaction
Published: 04.06.2018; Views: 187; Downloads: 45
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Poezija Franceta Prešerna v srednješolskih učbenikih in njena recepcijazoran Božič
, 2010, doctoral dissertation
Abstract: Disertacija obravnava dva problemska sklopa: poezijo Franceta Prešerna v srednješolskih učbenikih za pouk književnosti od leta 1850 do leta 2010 ter recepcijo Prešernovih pesmi skozi čas in v sodobnosti. V prvem problemskem sklopu razkriva dejavnike in postopke šolske kanonizacije Prešerna ter razvoj didaktizacije beril v smeri učne knjige za samostojno delo z besedilom. V drugem problemskem sklopu raziskuje posebnosti pri razumevanju, doživljanju in vrednotenju Prešernovih pesmi ter načine za njihovo čimbolj učinkovito šolsko predstavitev.
Glede na to, da se je kanonizacija Prešerna začela že za časa njegovega življenja, je bil seveda vseh 160 let kanonski avtor, vendar pa kljub nenehnemu označevanju Prešerna za "največjega slovenskega pesnika" po številu besedilnih enot v berilih ni bil vedno prvi avtor, pač pa so ga večkrat prehitevali mlajši književniki, ki so bolj ustrezali trenutnim afinitetam sestavljavcev srednješolskih beril. Predvsem v drugi polovici 19. stoletja in tudi kasneje je bil zaradi negativnega odnosa šolskih oblasti do ljubezenske poezije izbor Prešernovih pesmi praviloma manj primeren (na nižji stopnji je bil največkrat objavljen sonet Memento mori, na višji stopnji pa cikel Soneti nesreče), nekatere ljubezenske pesmi pa so celo cenzurirali.
Od druge polovice 19. stoletja do danes se je razvijal didaktični instrumentarij srednješolskih beril (najprej so začeli dodajati stvarne opombe, kasneje pa še vprašanja in naloge za dejavno razčlembo leposlovnega besedila), vendar so še zlasti v obdobju po drugi svetovni vojni nekateri sestavi beril didaktično pomenili izrazito nazadovanje. Od sedemdesetih let dalje berila vključujejo tako literarnovedne vsebine kot vprašanja in naloge, sodobna berila pa se praviloma (izjema je npr. antološki sestav Krakar 2003) razvijajo v smeri vezanega berila z relativno majhno ponudbo leposlovnih besedil.
Zgodovinski pregled je pokazal, da so imeli težave z razumevanjem Prešernovih pesmi že njegovi sodobniki, kasneje pa tudi izobraženci, celo poznavalci Prešerna. Empirični preizkusi med slovenskimi srednješolci potrjujejo visoko recepcijsko zahtevnost Prešernove poezije. Nekatere osrednje Prešernove pesmi (npr. Krst pri Savici) zahtevajo izjemno gostoto dodanih opomb, zato je najustreznejše didaktično sredstvo za boljše razumevanje in doživljanje oblikovno in slogovno zahtevne klasične poezije dodana prozifikacija, kar so v celoti potrdili tudi rezultati pedagoškega eksperimenta.
Keywords: poezija, Prešeren, didaktika, recepcija, kanonizacija, srednja šola, berilo, kanonizacija, prozifikacija, pedagoški eksperiment
Published: 12.05.2016; Views: 1176; Downloads: 22
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On non-simultaneous phasesFranc Marušič
, 2005, doctoral dissertation
Keywords: jezikoslovje, slovenščina, skladnja, teorija faz, minimalizem, rekonstrukcija, dvig kvantifikatorjev, elektronski vir
Published: 15.10.2013; Views: 2063; Downloads: 9
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