Dissertations / Theses on the topic 'Two-dimensional gas chromatography'

Separation technologies have occupied a central role in the current practices of analytical methods used for drug analysis today. As the emphasis in contemporary drug analysis shifts towards ultra-trace concentrations, the contribution from unwanted matrix interferences takes on greater significance. In order to single out a trace substance with confidence from a rapidly expanding list of drug compounds (and their metabolites) in real complex specimens, analytical technologies must evolve to keep up with such trends. Today, the task of unambiguous identification in forensic toxicology still relies heavily upon chromatographic methods based on mass spectrometric detection, in particular GC-MS in electron ionisation (EI) mode. Although the combined informing power of (EI) GC-MS has served faithfully in a myriad of drug application studies to date, we may ask if (EI) GC-MS will remain competitive in meeting the impending needs of ultra-trace drug analysis in the fut ure? To what extent of reliability can sample clean-up strategies be used in ultra-trace analysis without risking the loss of important analytes of interest? The increasing use of tandem mass spectrometry with one-dimensional (1D) chromatographic techniques (e.g. GC-MS/MS) at its simplest, considers that single-column chromatographic analysis with mass spectrometry alone is not sufficient in providing unambiguous confirmation of the identity of any given peak, particularly when there are peak-overlap. Where the mass spectra of the individual overlapping peaks are highly similar, confounding interpretation of their identities may arise. By introducing an additional resolution element in the chromatographic domain of a 1D chromatographic system, the informing power of the analytical system can also be effectively raised by the boost in resolving power from two chromatographic elements. Thus this thesis sets out to address the analytical challenges of modern drug analysis through the application of high resolut ion comprehensive two-dimensional gas chromatography (GC„eGC) to a series of representative drug studies of relevance to forensic sciences.

Thesis (MSc)--Stellenbosch University, 2011.
ENGLISH ABSTRACT: This study focused on the potential of comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry (GC×GC-TOF-MS) for the improved analysis of volatile wine constituents. Solid phase microextraction (SPME) in combination with GC×GC-TOF-MS was successfully used for the detailed investigation of the impact of three commercial Oenococcus oeni lactic acid bacteria (LAB) strains on the volatile composition of Pinotage wines subjected to malolactic fermentation (MLF). Due to increased separation power and enhanced sensitivity obtained by using two orthogonal separations coupled with the structural information provided by deconvoluted TOF-MS spectra, GC×GC-TOF-MS allowed for the identification and semi-quantitative analysis of much larger numbers of compounds compared to previous studies applying one-dimensional gas chromatography. The combination of univariate and multivariate statistical assessment was used as a powerful tool for data interpretation. The obtained results contribute significantly to the understanding of the impact of MLF on the volatile composition of Pinotage wine Some compounds have been linked to MLF for the first time. Moreover, the impact of these commercial starter cultures on the composition of volatile sulfur and nitrogen compounds in the same wines was studied by one-dimensional gas chromatographic methods with headspace injection and solid supported liquid-liquid extraction together with sulfur selective detection and tandem mass spectrometry. This study demonstrated also for the time, the impact of MLF on the composition of volatile sulfur and nitrogen compounds in Pinotage wine. GC×GC-TOF-MS was further used for the evaluation of the suitability of a new phase for stir bar sorptive extraction (SBSE) analysis of wine volatiles. Despite instrumental complications, beneficial extraction properties of the new stir bar phase for especially more polar compounds could be demonstrated. In addition, the extraction ability of this novel phase was evaluated for the analysis of selected thiazoles in wine using heart-cutting two dimensional gas chromatography in combination with nitrogen selective detection. Advantageous extraction performance of the new stir bar phase compared to a conventional polydimethylsiloxane (PDMS) phase for the determined thiazoles was demonstrated.
AFRIKAANSE OPSOMMING: Hierdie studie het gefokus daarop om die potensiaal van omvattende tweedimensionele gaschromatografie gekombineer met vlugtyd massaspektrometrie (GC×GC-TOF-MS) vir die verbeterde analise van vlugtige wynkomponente te ondersoek. Soliede fase mikro-ekstraksie (SPME) in kombinasie met GC×GC TOF MS is met sukses aangewend vir ‘n ondersoek na die impak van drie kommersiële Oenococcus oeni melksuur bakteria (LAB) rasse op die samestelling van die vlugtige fraksie van Pinotage wyne wat appelmelksuurgisting (AMG) ondergaan het. As gevolg van die verbeterde skeidingsvermoë en die verhoogte sensitiwiteit wat verkry word deur twee ortogonale skeidings te kombineer, tesame met die inligting aangaande die molekulêre struktuur wat die die gedekonvoleerde TOF massaspektra verskaf, maak GC×GC-TOF-MS die identifikasie en semi-kwantitatiewe analise van aansienlik meer komponente, in vergelyking met die gebruik van een-dimensionele gaschromatografie, moontlik. Die kombinasie van monoveranderlike asook multiveranderlike statistiese evaluering is gebruik as ‘n kragtige tegniek vir data interpretasie. Die resultate wat verkry is dra tot ‘n groot mate by tot die ontrafeling en begrip aangaande die impak wat AMG op die samestelling van vlugtige komponente in Pinotage wyn het. Daar word ook vir die eerste keer aangetoon dat somminge komponente verband te hou met AMG. Aanvullend hiertoe is die impak wat hierdie kommersiële kulture (wat gebruik word om fermentasie te inisieer) op die voorkoms van swawel en stikstof bevattende vlugtige komponente het bestudeer deur gebruik te maak van een-dimensionele gaschromatografiese metodes met ‘headspace’ inspuiting en vloeistof-voeistof ekstraksie tesame met swawel en stikstof selektiewe deteksie en tandem massaspektrometrie. Hierdie ondersoek werp lig, ook vir die eerste keer, op die samestelling van vlugtige swawel en stikstof bevattende komponente in Pinotage wyn. GC×GC-TOF-MS is ook gebruik vir die evalueering van die toepaslikheid van ‘n nuwe stasionêre fase vir gebruik met roerstaaf sorptiewe ekstraksie (SBSE) vir die analisering van vlugtige komponente in wyn. Ten spyte van instrumentele komplikasies, is die voordele wat hierdie nuwe fase vir die ekstraksie van vernaamlik meer polêre komponete aangetoon. Vervolgens is die ekstraksievermoë van hierdie nuwe fase vir die analise van sekere tiasole in wyn met ‘heart-cutting’ twedimensionaly gaschromatografie in kombinasie met stikstof-selektiewe deteksie gedemonstreer. Verbeterde ekstraksie van die nuwe roerstaaf fase vir die analise van tiasole, in vergelyking met ‘n tradisionele polydimethylsiloxane (PDMS) fase is voorts aangetoon.

Volatile organic compounds (VOCs) are key precursors to ozone and particulate matter, two of the most important air pollutants. Air quality interventions have successfully reduced the release of short chain VOCs in urban areas. The increased use of diesel vehicles has created an increase in the direct emission of longer chain VOCs. However, these compounds are not considered as part of air quality strategies and there are few atmospheric measurements of them to date. This thesis details continuous measurements of VOCs in London, a developed megacity, using comprehensive two dimensional gas chromatography. Analysis of this large suite of VOC measurements have shown that the higher carbon number species emitted from diesel vehicles can dominate gas phase reactive carbon in cities with a significant diesel fleet. Comparison of these real-world observations with emissions inventories has highlighted that there is a significant under prediction of the emissions of higher carbon number species. This presents a considerable policy challenge; the focus must now switch to VOCs released from diesel as this vehicle type is increasingly replacing gasoline world-wide. Further analysis of the London data has provided evidence of both anthropogenic and biogenic emission sources. The measurement of the higher carbon number species has allowed for OH reactivity to be more accurately modelled. Detailed analysis of the ethanol observations provided direct evidence that the use of bio-ethanol blended gasoline in the UK is having an impact on the composition of the atmosphere. The combination of heart-cut and comprehensive two dimensional gas chromatography into a single instrument has made the measurement of both small and large chain VOCs possible. This instrument compares well to existing instrumentation and when deployed to a rural location (Bachok, Malaysia) provided hourly time-resolved measurements of C5-C13 VOCs.

The use of short columns, higher carrier gas velocity and fast temperature programs in Comprehensive Two-Dimensional Gas Chromatography coupled to Time-of- Flight Mass Spectrometry (GC x GC/TOFMS) technique is expected to increase the speed of analysis up to several orders of magnitude when compared to conventional gas chromatography (GC) or gas chromatography/mass spectrometry (GC/MS). A systematic evaluation of the GC x GC/TOFMS configuration for high-speed applications has received little attention in the literature. The feasibility of High Speed Comprehensive Two-Dimensional Gas Chromatography coupled to Mass Spectrometry (High speed GC x GC/MS) for complex mixtures has been investigated in this thesis. A particular focus was placed on comparing conventional scanning quadrupole mass spectrometry (qMS) with a newly available non-scanning time-of-flight instruments (TOFMS). Experiments were carried out using GC/qMS, GC x GC/qMS, GC/TOFMS and GC x GC/TOFMS both in normal (slow) and fast temperature rates coupled with high frequency modulation in GC x GC. Initially a complex mixture consists of 24 semivolatile compounds was used as the analyte for the above purpose. In the initial experiments parameters like acquisition rate and duty cycle for qMS were determined to evaluate the effectiveness of the instrument for fast analysis. The practical duty cycle value obtained for the qMS was only about 18 % for single ion and one compound at a dwell time of 10 ms in SIM mode. In both high-speed GC/qMS and high-speed GC x GC/qMS techniques only about 40 % of the components in the complex mixture were found to be well separated. The acquisition rate of scanning instruments like qMS is incompatible for fast eluting peaks in high speed GC. TOFMS that has an acquisition rate of several hundred spectra per second offer the potential to define the fast GC peaks accurately. The high quality spectra from TOFMS also enable deconvolution of coeluting peaks in the complex mixtures. The advantage of the automated spectral deconvolution is demonstrated for the identification of the coeluting peaks in the complex mixtures. Coelution of peaks is also observed with highspeed GC/TOFMS technique. The high-speed GC x GC/TOFMS was also tested with two different analyte system ??? A pesticide mixture and platformate (an aromatic mixture) to evaluate the suitability for high-speed analysis of complex mixtures. A poor resolution was observed for the pesticide mixture in the two-dimensional plane and it appeared, as there was nearly no orthogonal separation in the second dimension. The platformate mixture displayed a better two-dimensional separation. Chromatographic peak resolution is not really a primary requirement for locating and identifying the coeluting compounds in high-speed GC x GC/TOFMS technique. However, it was observed that the high-speed GC x GC/TOFMS too faced problem to unscramble the mass spectra of those compounds with similar structure and sharing the same unique masses.

There are 209 PCB congeners, 136 of which have been found in technical PCB mixtures and hence may be found in the environment as a result of either intentional or unintentional release. The identification and quantification of the congeners are difficult due to analytical bias from coeluting PCBs and other persistent organic pollutants. Among the 209 possible PCB congeners, 19 tri- and tetra-ortho chlorinated congeners exist in stable atropisomeric conformations. The racemization barrier were determined for twelve of the nineteen atropisomers and was found to be between 176-185 kJ × mol-1 and ca. 250 kJ × mol-1 for tri- and tetra-ortho PCB, respectively. Further, a buttressing effect of 6.4 kJ × mol-1 was observed for congeners with vicinal ortho-meta chlorines. Comprehensive two-dimensional gas chromatography (GC×GC) was used to analyze the atropisomers and other PCBs. A Longitudinally Modulated Cryogenic System (LMCS) was used with liquid CO2 as cryogen. The LMCS was optimized for semi-volatile organic substances, primarily PCBs. The trap temperature was shown to be an important factor for the trapping and desorption efficiency, as was the thermal mass of the column used in the modulator region. A number of column sets were tested and the separation efficiency, congener resolution and analysis time was evaluated. Good separation of non- and mono-ortho PCBs and “bulk” PCBs (in a technical PCB) was obtained within 8 min using a smectic liquid crystal column (LC50) as the first and a nonpolar column as the second dimension column. Using a second column, an efficient nonpolar (DB-XLB) column, which separates many PCB congeners, were combined with a polar (cyanopropyl) or shape selective (LC50) second dimension column. As a maximum, 181 of the 209 congeners and 126 of the 136 Aroclor PCBs were resolved. The seven frequently measured PCBs (PCBs 28, 52, 101, 118, 138, 153 and 180) and all WHO-PCBs were separated from all other Aroclor PCBs. Chiral PCBs are released into the environment as racemic mixtures. However, organisms have been shown to enantiomerically enrich many of the atropisomers, suggesting that enantioselective biotransformations occur. Non-racemic PCB enrichment has also been seen in mammalians including humans, which is of particular concern because of the potential health risk. An analytical procedure were therefore developed and used to determine the levels of atropisomeric PCBs, planar-PCBs (WHO-PCBs) and total PCBs in seals with different health status. GC×GC was used to separate the target PCBs from other PCBs and potential interferences. A chiral column (permethylated â-cyclodextrin) was used in combination with a polar or shape selective column and enantiomeric fractions (EFs) were determined for five atropisomeric PCBs, i.e. CBs 91, 95, 132, 149 and 174. Some atropisomers had EF that deviated largely from racemic. The deviation was larger in liver than blubber, indicating enantioselective metabolism. However, there was no selective passage of the studied atropisomeric PCBs across placenta and no selective blood-brain barrier. Similarly, no correlation between EFs and health status was observed, although there was a correlation between total PCBs and health status.

AbstractComprehensive two-dimensional gas chromatography is a multidimensional separation technique. A sample is separated by two properties on two different columns, typically by carbon number and polarity. The two columns are connected by a modulator. The modulator is responsible for collection of three to four fractions of each 1st column separation peak, condensation of the fractions, and introducing them as a sharp narrow band onto the 2nd column. It is a continuous process of condensation of succeeding 1st column fractions and transfer to the 2nd column. The individual separations are “sewn” together by the software to produce a two-dimensional chromatogram. The abscissa displays the carbon number separation and the ordinate axis show the separation of polar compounds.Pre-set parameters such as carrier gas, gas velocities, detector temperatures, and column set were kept on recommendation by the installation contractors. Method development and optimisation was performed by exploring injection volume, oven temperature programs, and modulator time parameters. Hydrocarbon standards and petroleum fractions were analysed for determining the optimal parameter values. The result was two methods, one recommended for atmospheric gas oil (AGO) analyses and another for vacuum gas oil (VGO) analyses. Injection volumes of 0.015 to 0.002 µL gave low risk of column overload while still maintaining the abundance of compounds of low concentration. Temperature programmes of constant ramps gave good separation. A compromise between excellent separation and time of analysis resulted in using temperature ramps of 4.5 oC/min with a start and final temperature of 50 oC and 340 oC, respectively, for AGOs, and 3.5 oC/min with a start and final temperature of 150 oC and 340 oC, respectively for VGOs. A modulation time of 8000 ms and a hot jet duration of 500 ms proved to give good correlation between the 1st separation’s peak widths and the time needed for 2nd dimension separation. Straight run and processed petroleum fractions were analysed by the optimised methods. Constructed templates for dividing the sample’s polarity distribution into groups gave a distribution of volume response of all the compounds within the defined groups. The hydrocarbon analyses of the petroleum fractions were straight forward; volume responses were directly proportional to weight percent of the sample. The hydrocarbon standards gave approximately the same response factor. The same did not apply for sulphur analysis. The standards’ responses were not very reproducible, and the response factors were not similar for the polarity classes. Identification of sulphur compounds in AGOs and VGOs is possible although quantification is not recommended at the present. The methods showed to give good separation of both AGOs and VGOs. Although further optimisation especially of sulphur analysis, is highly encouraged.

Includes abstract.
Includes bibliographical references (leaves 158-178).
Fischer-Tropsch synthesis is a process that catalytically converts hydrogen and carbon monoxide into a large variety of hydrocarbons and oxygenated products. Over the years many researchers have attempted to describe the full product spectrum (ranging from C1 to C100+) but due to the complexity of the product and shortcomings of certain analytical techniques (or equipment) most researchers were only able to construct product distributions from extrapolations of data recorded from analysis of the C1 to C5 fraction of the Fischer-Tropsch product. With recent advances in analytical technology and the development of comprehensive two-dimensional gas chromatography (GCxGC) it may now be possible to analyze the complex Fischer-Tropsch products in a relatively short time while delivering good separation of even minor compounds such as oxygenates and branched compounds. The aim of this study was to investigate if two-dimensional gas chromatography (GCxGC) really results in improved separation and identification of compounds in the complex Fischer-Tropsch product spectrum and will lead to a more complete product distribution especially of the minor compounds such as branched hydrocarbons, ketones, aldehydes and acids. For this study GCxGC equipment, supplied by Zoex Corporation, was connected to a micro slurry phase reactor system to provide for both on-line gas analysis as well as off-line product analysis. GCxGC methods were developed to analyze the hot tail gas and oil products from Fischer-Tropsch synthesis. Thereafter a test sample (C6 to C30 oil product from Fischer- Tropsch synthesis process) was injected several times into both the GCxGC and 1D GC systems. The purpose of this was to compare the detection ability and accuracy of the two instruments.

The use of comprehensive two-dimensional gas chromatography (GC x GC) for the analysis of volatile organic compounds in the atmosphere will be presented. Three different systems have been used and developed each suited to a particular aspect of atmospheric VOC analysis. A method for the offline analysis of whole air samples using a commercial GC x GC system coupled to a time-of-flight mass spectrometer (TOFMS) has been developed and has been used during an aircraft campaign. Calibration and precision data for 60 hydrocarbon species are presented, with %RSD ranging from 0.56 to 14.78 % and LOD ranging from 0.01 - 0.61 pptv for a l L sample. The developed method was used during the RONOCO campaign and data for 39 hydrocarbon species re- ported. The system allowed for an investigation into the aromatic complexity of the samples and the effect of these aromatic species on atmospheric reactivity was also investigated. Further investigation into the VOC complexity in the atmosphere was also performed. The development of a portable valve based GC x GC system is also presented. The system uses total transfer methodology to allow for sensitive cryogen free modula- tion. Analysis of gasoline and tea tree was performed and the performance of the system was assessed using gasoline. The instrument was coupled to a thermal des- orption unit for the analysis of atmospheric VOCs and gave LODs of between 2-4 pptv for selected hydrocarbons in a l L sample. Further development of the system making it suitable for fieldwork is also presented. A third miniaturised GC x GC system has been developed, particularly suited for the analysis of biogenic VOCs. The system is compact, low power and cryogen free, with the large GC oven replaced with a novel direct column heating method. The small footprint, power and consumable requirement make this system suitable for deployment for analysis in remote locations.

Coal tars are complex mixtures of inorganic and organic compounds, which are dominated by PAHs and were produced as a by-product of the former manufactured gas industry. Forensic analysis of coal tar samples was carried out using two dimensional gas chromatography coupled with time-of-flight mass spectrometry (GCxGC-TOFMS). This thesis first presents the application of existing multivariate statistical models developed using UK tar samples to tar samples from the USA as well as identifying 947 individual compounds present within the tars. This has important implications as this study demonstrated that statistical methods developed using UK tar samples can be successfully applied to non-UK tars. The thesis then presents the application of post extraction derivitisation to a creosote samples allowing for the detection of 255 compounds, the majority of which would not be detected without derivitisation. The analysis also detected 1505 individual compounds within the Creosote and provides the most comprehensive list of compounds detected within Creosote that has been produced. The analysis was also able to suggest the production process for the tar from which the Creosote was distilled. The use of both derivitisation and GCxGC were vital in providing this forensic information. The thesis then presents the analysis of 16 tar samples using GCxGC and post extraction derivitisation and the production of a database of 2373 individual compounds detected within the tar samples. The study showed that 163 individual compounds were present within all tar samples regardless of the production processused. This has important implications as environmental assessment usually focuses on a limited number of compounds, which could be expanded using the database presented within this thesis. Finally the thesis presented a study that analysed a sample of Pintsch Gas tar, which has never previously been analysed using a mass spectrometer and produced a unique dataset.

Polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs), two groups of struc-turally related chlorinated aromatic hydrocarbons, generally referred to as “dioxins” are of great concern due to their extreme toxicity and presence in all compartments of the environment. Since they occur at very low levels, their analysis is complex and challenging, and there is a need for cost-efficient, reliable and rapid analytical alternatives to the expensive methods in-volving use of gas chromatography high-resolution mass spectrometry (GC-HRMS). It is im-portant to routinely monitor food and feed items to detect contaminations at an early stage. For the regulation of dioxins and dioxin-like PCBs in food and feed according to current legis-lation, large numbers of samples have to be analysed. Furthermore, soils at many industrial sites are also contaminated with dioxins and need remediation. In order to optimize the cost-efficiency of reclamation activities it is important to acquire information about the levels and distribution of dioxins in the contaminated areas. The aim of the studies underlying this thesis was to investigate the potential of comprehen-sive two-dimensional gas chromatography with a micro-electron capture detector (GC × GC-µECD) as a cost-effective method for analysing dioxins and dioxin-like PCBs in food, feed, fly ash and contaminated soils. Quantification studies of dioxins and dioxin-like PCBs were con-ducted and results were compared with GC-HRMS reference data. Generally, there was good agreement between both the congener-specific results and data expressed as total toxic equiva-lents (TEQs). The developed GC × GC-µECD method meets the European Community (EC) requirements for screening methods for control of dioxins and dioxin-like PCBs in food and feed. The presented results also indicate that GC × GC-µECD has potential to be used as a routine method for the congener-specific analysis of 2,3,7,8-PCDD/Fs and dioxin-like PCBs in matrices such as food and feed, fly ash and soil. However, to fully exploit the potential of the GC × GC-µECD technique, it should be combined with a fast and cost-efficient sample preparation procedure. Therefore, a number of certified reference materials (CRMs) were extracted using a new shape-selective pressurized liquid extraction technique with integrated carbon fractionation (PLE-C), and the purified extracts were analysed for PCDD/Fs using GC × GC-µECD. The results compared well with the certified values of a fly ash and a sandy soil CRM, but they were much too high for a com-plex clay soil CRM. It was concluded that this combination of techniques was very promising for screening ash and highly permeable soils. Further assessments and method revisions are still required before GC × GC-µECD can be used on a routine basis, and available software packages need to be refined in order to accelerate the data-handling procedures, which currently restrict the sample throughput.

Orientador: Fabio Augusto
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica
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Resumo: O projeto objetivou estudar os fundamentos que governam um sistema GC×GC, por meio da construção do principal componente de tal sistema, o modulador, sua adaptação a um cromatógrafo a gás comercial e posterior aplicação. O ponto de partida do desenvolvimento instrumental foi a projeção e a construção de um modulador de duplo jato com dois cabeçotes de aço e dióxido de carbono líquido como agente crioscópico. Numa segunda etapa os dois cabeçotes foram substituídos por peça única, utilizando-se ainda o mesmo fluido para resfriamento. Posteriormente o modelo em peça única foi alimentado por nitrogênio líquido. Ambos os sistemas moduladores foram controlados por softwares escritos em laboratório. O primeiro, utilizado em testes iniciais, foi escrito em Visual Basic 3. Em um segundo momento, foi escrito simultaneamente ao desenvolvimento do modulador um software exclusivo para o controle da modulação e registro dos dados. Durante todo o trabalho, algumas aplicações foram feitas para testar o desempenho do sistema. Foram realizadas separações GC×GC principalmente de amostras de gasolina Tipo C. Os resultados obtidos foram extremamente promissores uma vez que ficou demonstrada a aplicabilidade de um sistema modulador para GC×GC utilizandose um forno secundário, externo ao forno do cromatógrafo
Abstract: The aim of this project was to investigate the basic concepts concerning a GC×GC system by adjusting the main system component, the modulator, to a commercial gas chromatograph and its subsequent application. The instrumental development starting point was the projection and construction of a two piece dual jet modulator with carbon dioxide as coolant. In a second stage the two nozzles were replaced by a single piece and the same coolant was used. In sequence nitrogen gas cooled by liquid nitrogen was used as modulating agent with the single piece design. For system control and data acquisition two homemade software were used. The first was written in Visual Basic 3. The second was developed simultaneously to the modulator and is an exclusively applied version. During project development some applications were showed to evaluate system performance. GC×GC separations were performed mainly by using Brazilian gasoline C type. The results obtained are very promising once was demonstrated the applicability of a modulator system using a secondary oven external to the chromatograph oven
Doutorado
Quimica Analitica
Doutor em Ciências

The oxidation of volatile organic compounds (VOCs) plays a role in both regional and global air quality through the formation of secondary organic aerosols (SOA). More than 1000TgC/yr of non-methane VOCs are emitted from biogenic sources (significantly greater than from anthropogenic sources). Despite this magnitude and potential importance for air quality, the body of knowledge around the identities, quantities and oxidation processes of these compounds is still incomplete (e.g., Goldstein & Galbally, 2007; Robinson et al., 2009). Two-dimensional gas chromatography paired with time-of-flight mass spectrometry (GC×GC/TOFMS) is a powerful analytical technique which is explored here for its role in better characterizing biogenic VOCs (BVOCs) and thus SOA precursors. This work presents measurements of BVOCs collected during two field campaigns and analyzed using GC×GC/TOFMS. The first campaign, the Bio-hydro-atmosphere Interactions of Energy, Aerosols, Carbon, H2O, Organics & Nitrogen - Rocky Mountain Biogenic Aerosol Study (BEACHON-RoMBAS), took place in a Ponderosa pine forest in Colorado. The second campaign, Particle Investigations at a Northern Ozarks Tower: NOx, Oxidant, Isoprene Research (PINOT NOIR) Study, was conducted in the Ozark region of Missouri. Tens to hundreds of BVOCs were quantified in each set of samples, including primary emissions, atmospheric oxidation products, stress indicators and semi-volatile leaf surface compounds. These findings highlight that there is a largely uncharacterized diversity of BVOCs in ambient samples. Our findings demonstrate that GC×GC can distinguish between compounds with the same molecular weight and similar structures, which have highly variable potentials for production of SOA (Lee et al., 2006). This work represents some of the first analysis of ambient BVOCs with this technology, which is anticipated to contribute greatly to characterization of atmospheric SOA precursors and ultimately, regional and global modeling of SOA and fine particulate matter.

Cette thèse a été conduite afin d’explorer le potentiel d’une nouvelle méthodologie utilisant la pyrolyse, la chromatographie gazeuse et la spectrométrie de masse pour la caractérisation et l’identification des fibres papetières utilisées dans la fabrication des papiers asiatiques traditionnels à partir de la caractérisation des métabolites de ces fibres. Cette méthodologie utilise un processus d’échantillonnage facilité nécessitant une très petite quantité d’échantillons (de l’ordre de quelques dizaines de µg). Après la pyrolyse des échantillons de papiers et la séparation chromatographique des composés formés, des distributions caractéristiques pour les métabolites des fibres papetières (considérant leur présence et leur intensité) ont été observées dans une région définie comme région d’intérêt dans les chromatogrammes: ces distributions se sont révélées spécifiques pour la caractérisation des papiers fabriqués à partir de différents types de fibres et ont été utilisées pour distinguer l’origine des différentes fibres papetières couramment utilisées dans la fabrication de papiers asiatiques traditionnels. Premièrement, les problèmes rencontrés dans l’étude des papiers faits à la main ont été présentés, comme l’origine de la fabrication du papier, l’incohérence de certains résultats dans l’identification des fibres (reportés dans différentes études scientifiques), les limites de la microscopie pour l’identification des fibres papetières d’origines botaniques similaires et les risques d’imprécision dans le référencement des échantillons. Tous ces problèmes montrent la nécessité d’explorer de nouvelles méthodes pour (1) améliorer la fiabilité de l’identification des fibres papetières des papiers asiatiques traditionnels, (2) valider et confirmer les résultats obtenus par l’analyse microscopique. À cette fin, dans un premier temps, des papiers asiatiques de référence ont été étudiés. Les résultats expérimentaux ont montré que les différentes fibres papetières utilisées pour la fabrication des papiers étudiés montraient des différences dans les distributions de leurs marqueurs spécifiques : par exemple, les fibres d’origine de la famille Moraceae montrent une distribution caractéristiques de composés triterpèniques alors que les fibres d’origine de la famille Thymelaeaceae montrent une distribution caractéristiques de composés de type stigmastanes. De leur côté, les fibres des plantes appartenant au groupe Ma montrent peu de métabolites caractéristiques. Les différences observées dans la distribution de ces métabolites ont été attestées par la comparaison entre distributions obtenues à partir des fibres végétales et celles des papiers faits à la main attestant de l’origine commune de ces métabolites issus des tissus végétaux d’origine. Ainsi, la méthodologie étudiée se révèle prometteuse en tant que méthode de chimiotaxonomie pour l’identification des fibres inconnues de papiers faits à la main. Avec les exemples d'applications fournies au cours du travail expérimental, le couplage de la pyrolyse, de la chromatographie en phase gazeuse et de la spectrométrie de masse (avec l’utilisation de la Py-GC/MS et de la Py-GCxGC/MS) a montré sa capacité à distinguer les fibres d'une même famille (qui peuvent présenter des caractéristiques similaires en microscopie) et peut ainsi constituer une méthode efficace d'identification des fibres et de validation des résultats d'identification obtenus par l'observation microscopique. Dans la présente thèse, les caractéristiques de la chromatographie gazeuse intégralement bidimensionnelle GCxGC, ses avantages pour les applications dans le domaine du patrimoine culturel et son apport potentiel pour le traitement des données 1D ont été discutées (...)
This study was conducted to explore a new methodology for handmade fiber characterizationand identification using pyrolysis, gas chromatography and mass spectrometry. It employseasy sampling process with minor quantity of samples required. After pyrolysis of handmadepapers, a featured metabolites distribution patterns (presence plus intensity) eluting in the defined region of interest (ROI) was observed to be characteristic for handmade papers of different material origins. The method utilizes these metabolites distribution patterns as markers to discriminate different fiber origins. Firstly, the problems encountered in the investigation of handmade papers were introduced such as the origin of papermaking, the inconsistency in the fiber identification results sometimes gained by different scholars, the limits of microscopy in identifying fibers from similar species and the likely imprecision of the reference sample labeling. All these problems showed the necessity to explore a new method in order to (i) make precise fiber identification of handmade papers and (ii) to validate or confirm the identification results obtained by microscopy. Then, modern reference handmade papers were firstly studied. The result revealed that different plant fibers used for papermaking have different marker distributions in the ROI, forinstance, the Moraceae family with a featured distribution of terpene compounds and theThymelaeaceae family with a featured distribution of stigmasta compounds. The fibers fromthe ma group usually revealed few compounds in the ROI. This metabolites difference in theROI was attested from the plant tissues with their similar distribution in handmade papers and plant raw fibers. Thus, the chosen methodology offers promise as a method of chemotaxonomy for unknown handmade paper fiber identification. With the examples ofapplications provided during the experimental work, the coupling of pyrolysis, gaschromatography and mass spectrometry (through the use of Py-GC/MS and Py-GCxGC/MS)showed its ability to distinguish fibers from the same plant family (that may present similar microscopic features) and thus, can constitutes an effective method for fiber identification as well as to validate the identification results of the microscopic observation. In the present thesis, the features of GCxGC and the benefits for cultural heritage applications and its help for the ID data treatment were discussed. The tested Py-GCxGC/MS methodology has been for the first time proposed in the cultural heritage field and it harbors the potential to promote the research in this domain, enhancing our capacity to handle small quantities of complex samples while providing an exhaustive response on its composition

Polychlorinated Biphenyls (PCBs) are a group of 209 ‘man-made’ chlorinated organic compounds that were widely used in the 20th century for a variety of industrial uses. PCBs were first commercially produced in the 1929 and were manufactured until the 1980s when their use was phased out due to environmental and human health risks. However, due to their widespread use and persistence they are ubiquitous in the environment and remain a contaminant of concern. The structural properties that determine the persistence of PCBs in humans were therefore elucidated by statistical analysis of data from the National Health and Nutrition Examination Survey (NHANES). PCBs with chlorine bonding in the 2,5-and 2,3,6- positions (and 2- in di- and tri-chlorinated congeners) were rapidly biotransformed and so can be classed as episodic congeners whereas PCBs with chlorine bonding in the 2,3,4-, 2,4,5-, 3,4,5-, and 2,3,4,5- positions were more resistant to biotransformation and can therefore be classed as steady state congeners. A fundamental requirement of using PCB signatures for environmental fingerprinting is an effective analytical method capable of producing high resolution signatures from biological samples. An extraction and clean-up method was developed that was successfully applied to different biological matrices (blood and tissues). A two dimensional gas chromatography with time of flight mass spectrometry (GCxGC-ToFMS) method was designed and optimised to provide a congener specific method capable of identifying 200 out of the 209 PCBs, with detection limits in human serum in the range of 1 to 10 ng g-1 lipid. The extraction and detection methods were used to determine the source of PCB contamination and age date exposure in workers at a transformer dismantling plant. A total of 84 different PCB congeners were identified in the sera of 30 workers with concentrations of the 7 indicator PCBs ranging from 1.2 - 39 μg g-1 lipid. Analysis of PCB signatures was able to distinguish recent from prolonged exposure and also identified an additional source of inhalation exposure in a subgroup of workers. Analysis of 12 different tissue types obtained from the common guillemot (Uria aalge) suggested a high degree of perpetuation between the PCB signature in different tissue types. This shows that comparative assessments can be undertaken between animals using different tissue types and that small (1 g) samples of blood can be used as a non-lethal sampling technique. The regional provenance of 25 wrecked Leach’s storm petrels (Oceanodroma leucorhoa) was also determined using PCB signatures. Results from GCxGC-ToFMS analysis revealed distinctively different PCB signatures in birds from Canada and Europe. The findings reported in this thesis enhance our understanding of PCB signatures in the environment and show how they can be used effectively to age date and identify the source of exposure in humans and animals.

Orientadores: Ronaldo Aloise Pilli, Fabio Augusto
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica
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Resumo: Nesta tese, foram construídos modelos quimiométricos a partir de cromatogramas de diferentes amostras obtidos por cromatografia gasosa bidimensional abrangente com detecção por ionização em chama (GC×GC-FID). Numa primeira aplicação, modelos foram desenvolvidos para identificar adulteração em gasolina. Para isso, uma amostra de gasolina não adulterada fornecida pela Agência Natural do Petróleo (ANP) foi utilizada na preparação de um conjunto de calibração e de um conjunto de validação de amostras de gasolinas adulteradas. Em seguida, o modelo construído foi utilizado para avaliar outras amostras de gasolinas e os resultados obtidos foram comparados com os resultados obtidos pela ANP. Em um segundo estudo com gasolinas, 51 amostras foram fornecidas pela ANP juntamente com os resultados dos testes físico-químicos utilizados pela agência na avaliação da qualidade da gasolina. O objetivo deste estudo foi a elaboração de modelos quimiométricos utilizando dados de GC×GC-FID para a previsão de parâmetros físico-químicos de gasolina. Os métodos de seleção de variáveis siPLS e algoritmo genético foram utilizados na preparação de modelos para a previsão da temperatura de destilação a 10, 50 e 90 % v/v de destilado e no ponto final da destilação. Um modelo PLS foi construído para a previsão da densidade das amostras de gasolinas. Em um terceiro estudo, utilizando PARAFAC e GC×GC-FID, foi realizada a diferenciação entre amostras de gasolina do Brasil e da Venezuela, devido ao intenso contrabando de gasolina existente da Venezuela para o Brasil. Em uma quarta aplicação, foi desenvolvido um algoritmo para seleção de intervalos em dados de ordem superior, o qual foi avaliado através da quantificação de alergênicos em perfumes. Por último, foi relatado o primeiro uso de MCR-ALS na construção de modelos quimiométricos utilizando-se dados obtidos por GC×GC-FID para quantificação de óleo essencial de alecrim em amostras complexas preparadas em laboratório
Abstract: In this thesis, some chemometric models were built using chromatograms of different kinds of samples obtained by comprehensive two-dimensional gas chromatography with flame ionization detection (GC×GC-FID). In the first application, models were developed to identify adulteration in commercial gasoline samples. A non-adulterated gasoline sample provided by the Agência Nacional do Petróleo (ANP) was used to build a calibration and a validation sample set of adulterated gasoline. Then, the built model was used to evaluate other gasoline samples and the obtained results were compared to the results obtained by ANP. In a second study with gasoline, 51 samples were supplied by ANP with the results of the physicochemical tests used by ANP to evaluate the quality of gasoline samples. The aim of this study was the elaboration of chemometric models using GC×GC-FID chromatograms to preview some of the physicochemical tests used by ANP. The variable selection siPLS and genetic algorithm methods were used to build multivariate models in order to preview the distillation temperatures of gasoline at 10, 50 and 90 % v/v of distillated and at the final point of the distillation. A PLS model was built to predict the density of the gasoline samples. In a third study, using PARAFAC and GC×GC-FID, it was performed the differentiation between Brazilian and Venezuelan gasoline, due to the intense smuggling of gasoline from Venezuela to Brazil. In a fourth application, an algorithm was developed for variable selection in multi-way data, which was evaluated by quantification of allergens in perfume. Finally, it was reported the first use of MCR-ALS algorithm to develop chemometric models using data obtained by GC×GC-FID to quantify rosemary essential oil in complex samples prepared in laboratory
Doutorado
Quimica Analitica
Doutor em Ciências

Orientador: Fabio Augusto
Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Química
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Resumo: Neste trabalho foram desenvolvidas metodologias para a análise de lipídios na forma de seus ésteres metílicos de ácido graxo (FAME) em diversas matrizes. Para isso foram empregadas e otimizadas de forma multivariada técnicas de derivatização dos lipídios por transesterificação por catálise básica, seu isolamento e concentração por Microextração em Fase Sólida (SPME) e análise por Cromatografia Gasosa Bidimensional Abrangente (GCxGC). A partir do perfil de extração foi escolhida a fibra comercial com revestimento de poli(dimetilsiloxano) (PDMS) com espessura de filme de 7 mm. O tempo de extração escolhido foi de 10 min pois atingiu a condição de sistema estacionário. Os parâmetros cromatográficos, tais como a programação de temperatura do forno, período de modulação e tipo de fase estacionária das colunas utilizadas no protótipo GCxGC, foram também otimizados. Além disso, durante o progresso deste trabalho, foi desenvolvido o primeiro protótipo brasileiro de GCxGC acoplado a um Espectrômetro de Massas com Analizador Quadrupolar rápido (GCxGCxqMS). Dentre os parâmetros operacionais do qMS, foram otimizados o intervalo de varredura do analizador quadrupolar e sua frequencia de aquisição. O sistema GCxGCxqMS foi empregado para a identificação dos FAME presentes nas amostras empregadas neste estudo, através do uso das informações obtidas pelos espectros de massas obtidos das amostras, com padrões analíticos e combinando o uso de índices de retenção (LTPRI) e da estrutura cromatográfica. Neste trabalho, devido ao incremento em detectabilidade e sensibilidade, foi possível observar diversos FAME comumente não detectados por Cromatografia Gasosa Convencional (GC), como FAME de comprimento ímpar de cadeia e a presença de agrupamento metila no carbono 2 (-br2). Além disso, em virtude da baixa abrangência do banco de dados disponíveis, foi possível, e necessário, criar um banco de dados com índices de retenção para análise de lipídios. Espera-se que estas metodologias desenvolvidas e os resultados apresentados possam ser empregados como ferramenta em estudos de lipidômica e aplicações correlatas
Abstract: The aim of this project was to develop alternative methodologies for lipidomic studies and related applications. A method for the analysis of lipids in different samples such as waxes, edible oils and cheek cells was developed. The isolation and concentration of the analytes was performed by Solid Phase Microextraction (SPME) combined with Comprehensive Two-dimensional Gas Chromatography (GCxGC). Firstly, the base-catalysed transesterification of the lipids into their respective fatty acid methyl esters (FAME) was optimized according to a multivariate model. Thus, the sample prepation was optimized considering the thickness of the coating and their respective extraction profiles. The choosen commercial coating was a poly(dimethylsiloxane) (PDMS) with 7 mm film thickness, the extraction time was 10 min where stationary state conditions were achieved. Secondly, chromatographic parameters were also optimized, such as the temperature programming, modulation period, the identity of the first and second dimensional columns used in GCxGC prototype. During the current study, the first brazillian prototype of a GCxGC coupled to a mass spectrometer with a rapid quadrupolar analyzer was developed (GCxGCxqMS). Also, the scan interval and acquisition frequency of the qMS were optimized. For the identification of the analytes similarity searches for mass spectra, analytical standards and the combination retention indexes with chromatographic structuration were used. Additionally it was possible to detected and identify FAME which couldn't be detected by conventional GC such as odd numbered chain length and branched FAME. Moreover, elution pattern of a branched FAME was reported, specifically the FAME with a methyl group in the carbon 2 (-br2). Because of the restricted content of the retention indexes and mass spectra database, the construction of a database with retention indexes for lipid analysis was built. Hence the ultimate goal of the current study is to present the potencial and applicability of SPME for lipidomic studies and related applications
Mestrado
Quimica Analitica
Mestre em Química

La chromatographie en phase gazeuse couplée à la spectrométrie de masse simple quadripolaire (GC-qMS) est la technique la plus utilisée dans l’exploration des acides organiques urinaires dans le cadre du diagnostic des aciduries organiques (AO). Cependant, après analyse par GC-MS de plus de 1000 échantillons d'urines recueillis auprès d'enfants atteints de troubles neurologiques, moins de 0,5% d'entre eux ont reçu un diagnostic positif. Ceci témoigne de l’urgence de rechercher de nouveaux biomarqueurs dans le domaine des troubles neurologiques associés à des erreurs innées du métabolisme (EIM).L'objectif principal de cette thèse a été de développer une nouvelle méthode d’analyse des échantillons urinaires par chromatographie bidimensionnelle couplée à la spectrométrie de masse (GCxGC-qMS) et d'évaluer son apport dans le profilage métabolique de ces échantillons. Par ailleurs, dans le cadre du diagnostic des AO, dans un objectif de simplification de la procédure de préparation de l’échantillon, nous avons évalué une autre approche d’analyse directe en temps réel couplée à la spectrométrie de masse « DART-MS » des acides organiques urinaires. La première étape de notre travail a été de développer un protocole de préparation de l’échantillon compatible avec une méthode de séparation bidimensionnelle. Puis, grâce à l’étude systématique des différentes conditions de séparation, nous avons développé une méthode par CGxCG-qMS parfaitement adaptée à la séparation des composés extraits.Après validation analytique, la méthode proposée a été ensuite évaluée par le profilage des composés organiques présents dans des échantillons urinaires prélevés chez des volontaires sains. Par rapport aux profils obtenus par une méthode par GC-qMS classique, optimisée, les profils chromatographiques obtenus par la méthode proposée présentent une sensibilité et une résolution nettement plus élevées. Ceci se traduit par la détection en GCxGC-qMS de nombreux composés supplémentaires, jusqu’à 92, selon les échantillons. Parmi les composés supplémentaires détectés et caractérisés, certains sont d’un intérêt diagnostic reconnu.Malgré les difficultés liées à l’étape de préparation de l’échantillon et au traitement des données, l’ensemble des résultats obtenus par la méthode proposée sur de nombreux échantillons urinaires a confirmé les potentialités et la nécessité d’utiliser la chromatographie bidimensionnelle dans le domaine de la découverte de candidats biomarqueurs. Il s’agit maintenant de poursuivre l’identification des composés supplémentaires détectés dans les échantillons urinaires étudiés et d’appliquer la méthode proposée à un plus grand nombre d’échantillons témoins et pathologiques afin d’essayer d’identifier des candidats biomarqueurs.Enfin, les essais préliminaires par la technique DART-MS, effectués dans un laboratoire de la Direction des Applications Militaires (DAM) du CEA, ont été axés sur la détection d’un panel d’acides organiques représentatifs des différentes anomalies rencontrées lors des AO. Malgré les effets de suppression ionique qui restent à éliminer, les résultats obtenus montrent que cette approche mérite d’être approfondie
Among inherited metabolic diseases, organic acidemia (OA) or organic aciduria is characterized by urinary excretion of abnormal amounts or types of organic acids. OA is mostly associated with genetic conditions resulting in a specific step of amino acid catabolism dysfunction. Such alterations can produce disease states that range from mild to lethal neurological involvement. Gas chromatography coupled to mass spectrometry (GC-MS) remains the most used analytical technique for detecting specific urinary organic metabolites related to OA. However, after analyzing more than 1,000 urine samples collected from children with neurological disorders, by this technique less than 0.5% of them were positive. Thus, the discovery of new biomarker candidates for other metabolic diseases is urgently needed. The main objective of this thesis was to evaluate a new method using two-dimensional gas chromatography coupled to mass spectrometry (GCxGC-MS) for the both qualitative and quantitative metabolic profiling of children’s urine. As compared to GC-MS, GCxGC-MS shows great resolution power and high peak capacity. For this purpose, we first developed a GCxGC-MS method with an appropriate sample preparation protocol for urinary organic acids profiling. Applied to urine samples of healthy children and children with neurological disorders, the proposed method showed high sensitivity and peak capacity thus opening new possibilities for the characterization of new biomarker candidates. For instance, by this technique, we were able to detect in urines samples more than ninety additional compounds, which are not detected by a conventional GC-MS method. However, taken together the obtained results show that while the GCxGC technique has unmatched power to separate compounds in a complex mixture, the sample preparation protocol remains a limiting step for the precise quantification of the detected compounds. On another hand, in order to reduce the sample preparation step, we evaluated the direct analysis in real time (DART) method for the urinary organic acids screening. The obtained results are very promising

This thesis is focused on the possibility of analysis of volatile organic compounds produced by monocytes during sepsis. Method of comprehensive two-dimensional gas chromatography with mass spectrometric detection was chosen for this purpose. Content of the first part was the optimization of the method of two-dimensional gas chromatography for the determination of volatile organic compounds. In this part were gradually adjusted parameters of the gas chromatography method to achieve the maximum efficiency. Further were adjusted conditions of samples preparation. Content of the second part was the usage of already optimized method for the analysis of the samples set of monocytes. Samples were subjected to the action of different inhibitors of the immune system and stimulators simulating bacterial or yeast infection. Based on this analysis were identified some compounds, which are produced by monocytes under condition simulating the infection.

Understanding the composition of atmospheric organic particulate matter (OPM) is essential for predicting its effects on climate, air quality, and health. However, the polar oxygenated fraction (PO-OPM), which includes a significant mass contribution from carboxylic acids, is difficult to speciate and quantitatively determine by current analytical methods such as gas chromatography-mass spectrometry (GC-MS). The method of chemical derivatization and two-dimensional GC with time of flight MS (GC×GC/TOF-MS) was examined in this study for its efficacy in: 1) quantifying a high percentage of the total organic carbon (TOC) mass of a sample containing PO-OPM; 2) quantitatively determining PO-OPM components including carboxylic acids at atmospherically relevant concentrations; and 3) tentatively identifying PO-OPM components. Two derivatization reagent systems were used in this study: BF₃/butanol for the butylation of carboxylic acids, aldehydes, and acidic ketones, and BSTFA for the trimethylsilylation (TMS) of carboxylic acids and alcohols. Three α-pinene ozonolysis OPM filter samples and a set of background filter samples were collected by collaborators in a University of California, Riverside environmental chamber. Derivatization/GC×GC TOF-MS was used to tentatively identify some previously unidentified α-pinene ozonolysis products, and also to show the characteristics of all oxidation products determined. Derivatization efficiencies as measured were 40-70% for most butyl derivatives, and 50-58% for most trimethylsilyl derivatives. A thermal optical method was used to measure the TOC on each filter, and a value of the quantifiable TOC mass using a gas chromatograph was calculated for each sample using GC×GC separation and the mass-sensitive response of a flame ionization detector (FID). The TOC quantified using TMS and GC×GC-FID (TMS/TOCGC×GC FID) accounted for 15-23% of the TOC measured by the thermal-optical method. Using TMS and GC×GC/TOF-MS, 8.85% of the thermal optical TOC was measured and 48.2% of the TMS/TOCGC×GC-FID was semi-quantified using a surrogate standard. The carboxylic acids tentatively identified using TMS and GC×GC/TOF-MS accounted for 8.28% of the TOC measured by thermal optical means. GC×GC TOF-MS chromatograms of derivatized analytes showed reduced peak tailing due in part to the lesser interactions of the derivatized analytes with the stationary phase of the chromatography column as compared to the chromatograms of underivatized samples. The improved peak shape made possible the greater separation, quantification, and identification of high polarity analytes. Limits of detection using derivatization and GC×GC/TOF-MS were μL injected for a series of C2-C6 di-acids, cis-pinonic acid, and dodecanoic acid using both butylation and TMS. Derivatization with GC×GC/TOF-MS was therefore effective for determining polar oxygenated compounds at low concentrations, for determining specific oxidation products not previously identified in OPM, and also for characterizing the probable functional groups and structures of α-pinene ozonolysis products.

Caractériser l’odeur humaine revêt un intérêt particulier en sciences forensiques. Les chiens peuvent retrouver une personne grâce à son odeur, mais leur incapacité à témoigner fait que l’information qu’ils apportent a peu de force probante devant les tribunaux. Ainsi, développer une stratégie analytique complète, du prélèvement des échantillons à l’analyse et au traitement des données, permettra d’appuyer les identifications effectuées par les chiens policiers. Après un état de l’art des différentes études qui se sont déjà penchées sur le sujet, nous nous pencherons sur l’utilisation des chiens en Gendarmerie et sur quelques aspects de la biométrie pour savoir dans quelle mesure l’odeur peut être utilisée comme une caractéristique biométrique. Nous avons développé des méthodes d’échantillonnage de l’odeur innovantes, simples et fiables, que ce soit directement sur le sujet ou indirectement sur les lieux du crime. Une attention particulière est portée aux échantillons réels et à leur conservation. L’aptitude des techniques analytiques à séparer et identifier les différents constituants de l’odeur humaine a été évaluée. Ainsi, nous avons développé une méthode d’analyse en chromatographie gazeuse bidimensionnelle intégrale couplée à la spectrométrie de masse. Les performances de plusieurs jeux de colonnes ont été comparées en considérant neuf des critères les plus utilisés dans la littérature. Enfin, nous avons constitué un panel représentatif sur lequel nous avons pu mettre en place les approches statistiques de comparaison des échantillons à des fins d’identification
Characterizing human odor is of particular interest in forensic science. Dogs can find a person using their odor-print, but cannot testify : the information they bring lacks probative value in the courts of justice. Thus, developing a whole analytical strategy, from sampling to analysis and data processing, may support the identification dogs provide. After reviewing the studies that have already worked on the subject, the use of dogs in the police force is developed and some aspects of biometrics are presented to know to if the odor can be used as a biometric trait. Innovative, simple and reliable methods were developed to sample odor, either directly on the subject or indirectly at the scene of the crime. Particular attention is paid to the real samples and their conservation. The ability of analytical techniques to separate and identify different constituents of human odor was assessed. Thus, we have developed an analytical method in two-dimensional gas chromatography integral coupled with mass spectrometry. The performance of several sets of columns was compared by considering nine of the criteria most used in the literature. Finally, we established a representative panel on which we were able to apply statistical approaches for the comparison of samples

Orientador: Fabio Augusto
Dissertação ( mestrado) - Universidade Estadual de Campinas, Instituto de Química
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Resumo: A carne de porco é a proteina mais consumida no mundo. Para ser consumida a carne necessita de um padrão de qualidade e um dos problemas que afeta esta qualidade é o surgimento de odores desagradáveis. Estes odores, conhecidos como boar taint, são causados principalmente pela presença de androstenona e escatol em níveis maiores que 1,00 mg g e 0,25 mg g, respectivamente, em gordura suína. Os métodos existentes para determinação destes compostos são dispendiosos e não geram resultados em tempos satisfatórios para análise de rotina. Neste trabalho foi proposto um novo método para determinação de androstenona e escatol em gordura suína que se baseia na utilização de microextração em fase sólida (SPME) e cromatografia gasosa bidimensional abrangente (GC xGC). Para isso foi realizada uma otimização univariada do tipo de fibra e agente saponificante e posteriormente uma otimização multivariada, envolvendo concentração de saponificante, tempo de saponificação, temperatura e tempo de extração. O método otimizado foi validado sob os parâmetros de limites de detecção e quantificação, linearidade, precisão e exatidão. Para o escatol é possível a quantificação de amostras com quantidades menores que as detectadas sensorialmente (0,25 mg g), com precisão e exatidão, mas para a androstenona, é possível apenas detectar a presença ou não deste analito. Após todo o desenvolvimento e validação do método, este foi aplicado em sete amostras de toucinho cedidas pelo ITAL
Abstract: Pork is the must consumed protein the world. For consumpted, the meat needs a quality standard and one of the problems concerning the quality of pork meat are off-flavours. These off-flavores, called boar taint are caused by the presence of two compounds, skatole and androstenone, in levels superior to 1.00 and 0.25 mg g, respectively in pig fat. In order to determine these flavors is necessary to know the concentrations of androstenone and skatole in pig fat. There are specific methods to analyse these compounds, but they spend a lot of time and require a great number of clean-up steps process. In this work, a faster and cleaner method was proposed to determine the presence of androstenone and skatole in pig fat, based in the solid phase microextraction (SPME) and comprehensive bidimensional gas chromatography (GC xGC). A univariate optimization considering the kind of fibre coating and hydrolysis agent was performed. Later a multivariate optimization was developed to determine the concentration of hydrolysis agent, time of saponification and temperature and time of extraction. The best conditions found for the method were used for its validation. The quantification and detection limits, linearity, accuracy and precision were determined. For skatole it is possible to quantify samples at smaller quantities than those found sensorially (0.25 mg g), with precision and accuracy, but for androstenone the method only detects its presence or absence in the analyte. After all the development and validation of the method, it was implemented with seven samples of pig fat, courtesy of ITAL (Instituto de Tecnologia de Alimentos)
Mestrado
Quimica Analitica
Mestra em Química

Orientador: Fabio Augusto
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Química
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Resumo: O presente trabalho é dedicado à aplicação de técnicas cromatográficas multidimensionais à problemas complexos de separação. No primeiro conjunto de estudos foram desenvolvidos métodos analíticos para investigação do metaboloma de plantas de interesse comercial. Para isso, foram utilizadas a cromatografia gasosa bidimensional abrangente (GC×GC-MS) e a cromatografia líquida de ultra eficiência (UHPLC-MS) acopladas à espectrometria de massas para aquisição do perfil metabólico de folhas de eucalipto. Logo, foram desenvolvidos modelos quimiométricos para interpretação dos dados e determinação de marcadores biológicos associados ao estresse biótico e ao fenômeno de resistência. Estes estudos permitiram aprimorar o entendimento do mecanismo de defesa de eucaliptos contra fitopatógenos. Em segundo plano, foram desenvolvidas fases estacionárias derivadas de líquidos iônicos (IL) para separação de compostos apolares por GC×GC. Para isso, foram sintetizados diversos IL derivados de fosfônio e imidazólio. A partir destes materiais foram preparadas colunas capilares pelo método estático de revestimento. Estas colunas de GC foram utilizadas na separação de analitos modelo (i.e., hidrocarbonetos alifáticos) por GC×GC. Estes ensaios visaram aprimorar o entendimento da relação entre as características estruturais dos IL e os mecanismos que governam a retenção de compostos apolares pelas fases derivadas de IL
Abstract: In the present dissertation, we discuss the application of multidimensional chromatographic techniques to solve complex problems. In the first chapter it is presented the development of chemometric strategies for data processing of metabolic data ¿ the current bottleneck of a metabolomics workflow. The case studies examined investigate the metabolome of eucalyptus leaves to address several growing concern in plant pathology, namely, prospection of orthogonal methods for early diagnosis diseases and selection of hybrids with specific phenotypes in genetic enhancement programs. To accomplish these goals, comprehensive two-dimensional gas chromatography coupled to mass spectrometry (GC×GC-MS) and ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) were examined to generate unbiased and reliable metabolic profiles. From these experiments, it was possible to improve our understanding of the defense mechanism of plants. The second chapter addresses the limited availability of stationary phases for multidimensional gas chromatography. In this work, we evaluated ionic liquids as stationary phases for gas-liquid chromatography. The model analytes were nonpolar aliphatic hydrocarbons due to the aggravated lack of highly selective and thermally stable GC columns for their separation. From these experiments, we ascertained the structure-selectivity relationship of ILs and were able to improve our understanding on the retention of nonpolar analytes by IL-based GC columns
Doutorado
Quimica Analitica
Doutor em Ciências

This master thesis deals with the potential influence of the waste dump Hrádek u Pacova and the wastewater treatment plant Brno – Modřice on surroundings using the optimized non-target screening method of organic pollutants in water and sediments. The experimental part is focused on the identification of organic compounds in samples of ground water from monitoring wells, the surface water from stream and pond and in sediment samples collected from streams in the vicinity of municipal waste dump. Organic compounds were also identified in water and sediment samples from the river Svratka near the municipal waste water treatment plant Modřice. The sequential liquid – liquid extraction was used for the isolation of organic compounds in water samples. Organic compounds in sediment samples were isolated by pressurized solvent extraction (PSE) with subsequent fractionation of the extract using column chromatography. Final analysis determination was performed using comprehensive orthogonal two-dimensional gas chromatography with mass spectrometric detection (GCxGC-TOF MS).

Conjugated Linoleic Acid (CLA) are a group of positional and geometric isomers of Linoleic Acid characterized by a carbon chain containing 18 carbon atoms and two double bonds, not in the classic position (cis), but conjugated from the carbons atoms 9, 10 or 11. Double bonds have different position in the carbon chain ([7,9], [8,10], [9,11], [10,12], [11,13] and [12,14]) and four different geometric distribution (cis/trans, trans/cis, cis/cis and trans/trans). In total 24 possible isomers are identify, but the two most present and often most identified are: C18:2cis9,trans11 (60-85% of the isomers identify in meat and >90% in milk) and C18:2trans10,cis12 isomers. Observing meat samples other isomers are commonly identified, as example, C18:2trans7,cis9 and C18:2trans11,cis13. In 1987, CLA have been discovered thanks to the research group of Dr. Pariza that studying some carcinogenic components in grilled meat, identify these molecules with anticancer activity, called Conjugated Linoleic Acid. After the discovery, they still receive many attention because of their biological activities and implication on human health. The biological effect are due to the separate actions of the main isomers (C18:2cis9,trans11 and C18:2trans10,cis12) and sometimes by the synergistic action of both. Also, minor isomers have biological effect, as examples, C18:2trans9,trans11 that inhibits platelet aggregation and has anti-proliferative effect and C18:2cis9,cis11 that showed anticancer effect. In animal model, CLA not only reduce initiation, promotion and progression steps of cancer development, but also reduce metastasis. Nevertheless, was show that CLA isomers are important because of they have effect on animal performance and the principal effects are: prevent chemically-induced tumors, protect against the catabolic effects of immune stimulation, improve feed efficiency, reduce excess body weight gain, reduce body fat, increase lean body mass and lower blood lipids. In human diet, the main sources of CLA are representing by ruminants products, in particular milk, dairy product and beef. The higher concentration of CLA in these products is linked to the presence of rumen that through microbial biohydrogenation (Butyrivibrio fibrisolvens) can transform Linoleic Acid to the major isomer C18:2cis9,trans11 (Rumenic Acid). Conjugated Linoleic Acid (CLA) can be analyze with different methods of lipid extraction and derivatization, but always after transformation in methyl ester derivatives of fatty acids (FAMEs) that is carried out in a simple reaction. This simple reaction becomes more complicated because of conjugated fatty acids are involved and the presence of conjugated double bond makes them unsuitable for the most common techniques employed for fatty acids analysis. The presence of double bonds may increase isomerization and epimerization of these bonds, increasing CLA with trans/trans configuration and reducing cis/trans or trans/cis configuration. For this reason is very important find a suitable method for lipid extraction (and the resulting determination of crude fat) and subsequently for the transformation in methyl ester derivatives of fatty acids (FAMEs). For this reason at the first part of my thesis was given a methodological approach in order to understand the differences between methods and which is the most suitable (Chapter 2 and Chapter 7). All the data used, belonging to a trial carried out at “Lucio Toniolo”, the Experimental Farm of the University of Padova in Legnaro (Padova, Italy). Animals used are a crossbreed between Belgian Blue bulls and Brown Swiss dairy cows fed with one of 3 experimental diets. Diets were composed by unifeed and differ depending on the rumen protected CLA supplementation (rpCLA), overall, three supplementation are available 0, 8 and 80 g/d/animal. The trial ended in March 2011 when animals were slaughtered outside the faculty in a slaughterhouse located in Pergine province of Trento (Trentino Alto Adige Region). Samples were collected and in particular three tissues were subjected to analysis for fatty acids profile and CLA content: muscle Longissimus Thoracis, Subcutaneous Fat and Liver. In the main Chapter (Chapter 2) tissues (Longissimus Thoracis, Subcutaneous Fat and Liver) were analyzed for determine fatty acids profile comparing three different methods of extraction (Folch (1957), Accelerated Solvent Extraction (ASE) and Jenkins (2010)) using as chromatography technique two dimensional GC (GC×GC). The purpose of this trial was identify among the different methods of extraction, one method that is able to identify CLA isomers without causing isomerization. Folch (1957) is one of the older and most used methods. It was born for analyze samples rich in phospholipids (as lipids of brain). It works at room temperature using a mixture of solvents composed by chloroform/methanol (2:1, v/v). Accelerated Solvent Extraction (ASE) was used with the purpose to compare a room temperature method with a method that works at high temperature and pressure (120°C and 20MPa) with the same mixture of solvents (chloroform/methanol, 2:1, v/v). This method, which have recently been introduced; reduce the use of solvent and saving time at work, giving results that were similar or better if compared with the conventional Folch (1957) extraction. On the contrary, it is expected to increase the isomerization of double bonds and the isomers with trans/trans configuration. The last method, Jenkins (2010), is a direct method, which reduces the length of the total procedure, saving time at work, reducing the sample amount, reducing the use of solvents, giving analysis less expensive and easier. The main characteristic is that lipid extraction step is avoided and fatty acids are extracted and trans-esterified in the same time. The statistical analysis was performed in four steps: resolution power and assessment of the number of undetected FA, study the main sources of variation, using Levene’s test explored the variances homogeneity for the main sources of variations and relationships between methods. The incidence of undetectable values on the total number of expected observation, which depend on the sensitivity of the method used, for liver, fat and muscle, ranged 0.04 to 0.08, 0.05 to 0.06, and 0.05 to 0.12, respectively, with incidences greater for the Jenkins method compared to the other two for liver and muscle samples but not for subcutaneous fat. In liver the highest incidence of null values with the Jenkins method was mainly observed for short chain FA (C8:0 and C10:0), whereas in the case of muscle the highest incidence of undetectable values was mainly observed for the C24:0, for two C20:1t unknown isomers, and for Ω3 (C20:3n3, C20:4n3, C22:5n6). Many sources of variation result high significant (P<0.001). Diet was significant in particular for the FA that are the main components of the supplementation (C18:0, C18:1cis9, C18:2cis9,trans11 and C18:2trans10,cis12). The results of the Levene’s test evidenced, except for diet, as the variances among levels within tissue, method or method×tissue were not homoscedastic for the large majority of the FA. The second part of the methodological approach is reported in Chapter 7 (Minor Chapter) and considers the effect of the chromatography technique on CLA isomers content. Despite the high resolution power of GC×GC this technique allows to identify all the FA and the CLA isomers recognized by internal standard. However, it is not able to identify the other isomers and for this reason is necessary to find a new method that allows a clear and complete identification of CLA. Silver Ion High Performance Liquid Chromotography (Ag+HPLC) is currently the most effective way to separate and quantitate individual isomers of CLA in beef. This part was performed with the contribution of the Leibniz Institute for Farm Animal Biology (Dummerstorf, Germany) and in particular the Muscle and Biology Growth Unit. Tissue analyze in this trial were Longissimus Thoracis, Subcutaneous Fat and Liver. Lipids were extracted using Folch (1957) and methylated using and acid-base catalysis. Data were analyzed considering as main sources of variation: diet, tissue and repetition. Tissue resulted always significant (P<0.001) with a tissue depending distribution of the isomers and a higher concentration in subcutaneous fat. Diet was significant for the main isomers (C18:2cis9,trans11 and C18:2trans10,cis12) because of constituents rpCLA supplementation. In the second part of my thesis are considered the effects that can modify FA profile, such as, breed, gender, diet, type of birth, age and tissue. Animals used belonging to four native alpine sheep breeds: Alpagota, Brogna, Foza and Lamon. All these breeds represent an important genetic resources and the purpose is created a program of conservation for increase the number of animals in these populations. For this reason Veneto Agricoltura created the experimental farm “Villiago” (Belluno province, Veneto Region, North Italy). Animals used for this research belongs to two flocks undergoing an in situ conservation program between “Lucio Toniolo” Experimental Farm of the University of Padova and the Experimental Farm of Veneto Agricoltura located in Villiago. In total 115 animals were used and reared in the period between December 2010 and July 2012. Animals are slaughter at different age and they are considered belonging to three different categories: 31 suckling lambs, 36 lambs, 24 heavy lambs and 24 ewes. The different trial were characterized by different diets: pasture (PAS), penned in the open barn and fed with hay, concentrate and supplemented with rumen protected Conjugated Linoleic Acid (rpCLA) product (CLA+) and penned in an open barn and fed with hay and concentrate (CLA-). In trials with “lambs” and “heavy lambs” animals were supplement with 8.0 g/d/animal of rpCLA. In “suckling lambs” and “ewes” animals were supplement with 4.0 g/d/animal of rpCLA and 12 g/d/animal of rpCLA, respectively. From the results, is possible observe that diet is highly significant (P<0.001) and in particular pasture. Fatty acids are statistically influenced by pasture and mainly fatty acids with odd chain and branched fatty acids (iso and anteiso) that tend to be lower in diet at pasture. The reason is that the diet can influence ruminal Ph and microorganisms that consequently change reactions and final products. Pasture is also important because it can influence the amount of long chain fatty acids (LC-PUFA), increasing Ω3 (respectively, PAS=2.70, CLA-=1.46 and CLA+=1.54), reducing Ω6 (respectively, PAS=3.76, CLA-=4.41 and CLA+=4.75) and Ω6/Ω3 ratio. CLA isomers content is higher in pasture than in diet with concentrate supplement with rpCLA (respectively, PAS=0.80, CLA-=0.56 and CLA+=0.71). In the other two trials characterized by CLA+ and CLA- diet was considered an important and significant effect. rpCLA supplementation reported significant effect (P<0.001 and P<0.01) for FA constituents of supplementation, such as C18:0 and C18:2trans10,cis12. In suckling lambs results were different from other trials, in particular for CLA isomers, such as, C18:2cis11,trans13 (P<0.01), C18:2cis11,cis13 (P<0.001) and C18:2cis9,cis11 (P<0.001). The reason is the different distribution is ewe’s milk that was used in suckling lambs diet and could have influence their fatty acids profile. Breed was not a significant effect, such as, gender and age at slaughter. Observing data about orthogonal contrast FA and CLA isomers distribution is tissue specific and there are many differences between lean tissues (muscle) and fat tissues. Liver has particular characteristics, in fact, its content of PUFA and particular Ω3 and CLA is higher than in the others tissues. Differences of this tissue are linked to its specific lipid composition and metabolic process which lead to the production and absorption of fatty acids. In Chapter 5 and Chapter 6 were evaluate effects that CLA supplementation can have on animal performance because of many research have been conduct in vitro, but other research are needed. In Chapter 5 was examined the effect of rpCLA supplementation in lactating ewes on their milk composition and Milk Coagulation Properties (MCPs) of sheep’s milk. Animal used in this trial are the same of the trial describe above and in particular of the trial call “suckling lambs” and “ewes”. This study allowed to know that rpCLA supplementation in sheep can change the composition and cheese-making properties of milk, as example, delaying gelation, slowing curd firming and accelerated syneresis and future studies are necessary to know the effect of CLA on cheese yield/ quality. rpCLA supplementation affect milk composition, reducing protein content, solid non-fat content, casein index and increasing SCS. rpCLA supplementation had negative effects on parameters of coagulation and curd firming. In Chapter 6 was evaluate the effect of breed and sex on growth rate, slaughter traits and meat quality traits of lambs of Alpagota, Brogna and Foza breeds. Animals used in this trial are the same of the trial describe above and in particular of the trial call “lambs”. These results are part of a bigger study which comprises also growth rate, slaughter traits and meat quality of the trial call “heavy lambs”, “suckling lambs” and “ewes”. Observing the results of this trial is possible obtain lamb carcasses and meat with valuable characteristics that can be exploited through typical products and food preparation in local markets and gastronomy, according to tradition. The valorisation of these productions can be an important tool for the in situ conservation of these breeds. As example, Slow Food organization has recognized “Agnello Alpagoto” (lambs of Alpagota breeds) as a Slow Food Presidium.
Dal 1987 anno in cui il Dott. Pariza e il suo gruppo di ricerca scoprirono i Coniugati dell’Acido Linoleico (CLA) molti sono stati gli studi che hanno cercato di definire le principali caratteristiche di queste molecole. Caratterizzati da un alto valore biologico gli furono attribuiti molti effetti benefici sulla salute umana, come l’effetto anticancerogeno, la riduzione del rischio di malattie cardiovascolari e la riduzione del rischio di sviluppo dell’aterosclerosi. In seguito gli fu attribuita anche importanza nel miglioramento delle performance animali, come l’aumento delle capacità di accrescimento, dell’efficienza alimentare e una riduzione della deposizione di grasso con conseguente aumento della massa magra. I Coniugati dell’Acido Linoleico (CLA) sono un gruppo d’isomeri geometrici e posizionali dell’Acido Linoleico caratterizzati da una catena di 18 atomi di carbonio contenente due doppi legami non in posizione classica (cis), ma coniugati dal carbonio 9, 10 o 11. I doppi legami possono presentare diversa disposizione spaziale dando origine a quattro diverse configurazioni: cis/trans, trans/cis, cis/cis e trans/trans. Secondo i carboni ai quali sono legati, possono avere diverse posizioni: ([7,9], [8,10], [9,11], [10,12], [11,13] e [12,14]) con un totale d’isomeri identificati pari a 24. I due più presenti e più identificati sono il C18:2cis9,trans11 (60-85% degli isomeri presenti nella carne e >90% nel latte) e il C18:2trans10,cis12. Altri isomeri molto presenti nella carne sono anche il C18:2trans7,cis9 e il C18:2trans11,cis13. Grazie alla capacità del rumine di produrre acidi grassi e in particolare CLA, si possono trovare soprattutto nei prodotti di origine animale (latte, prodotti lattiero caseari e carne). La presenza del doppio legame rende i CLA delle molecole complesse da identificare, perché può essere facilmente soggetto a fenomeni d’isomerizzazione o epimerizzazione che possono portare a un aumento delle forme di tipo trans/trans con conseguente riduzione delle forme cis/trans o trans/cis. Diversi studi hanno cercato di definire quale sia il metodo più adatto per l’estrazione del grasso (determinazione estratto etereo, EE) e per la successiva trasformazione in composti volatili, ovvero, esteri metilici degli acidi grassi (FAME). Per questo motivo parte della mia tesi è stata improntata su un approccio metodologico allo scopo di capire tra i tanti metodi quale fosse il più idoneo e che differenze i diversi metodi potessero avere. I dati contenuti nei contributi legati all’approccio metodologico sono stati raccolti da vitelloni maschi nati da un incrocio tra vacche da latte di razza Bruna e tori di razza Bianca Blu del Belgio. Questi animali sono stati allevati presso l’azienda sperimentale dell’Università degli Studi di Padova Lucio Toniolo. Durante tutta prova sono stati allevati in azienda e alimentati con una dieta a base di unifeed caratterizzata da tre diverse integrazioni di CLA rumino protetti (rpCLA): 0, 8 e 80 g di CLA al giorno per ogni animale. La prova è terminata con la macellazione avvenuta in un macello esterno all’Università situato a Pergine (Provincia di Trento, Trentino Alto Adige). I tessuti utilizzati per le analisi sono stati prelevati in macello ed in laboratorio di Qualità carne durante lo svolgimento delle analisi di qualità. I tessuti prelevati e studiati sono stati tre: muscolo Longissimus Thoracis, grasso sottocutaneo e fegato. Nel primo contributo (Chapter 2) i tessuti (Longissimus Thoracis, grasso sottocutaneo e fegato) sono stati analizzati allo scopo di determinare il profilo acidico, confrontando tre diversi metodi di estrazione del grasso (Folch (1957), Acellerated Solvent Extraction (ASE) e Jenkins (2010)) e utilizzando come tecnica cromatografica la Gas Cromatografia a due dimensioni (GCxGC). La scelta di eseguire un confronto metodologico è legata al fatto che secondo il metodo d’analisi utilizzato i risultati sono diversi. Di conseguenza, uno degli obiettivi era trovare un metodo che fosse in grado di salvaguardare i CLA senza provocare isomerizzazioni. A questo è dovuta la scelta del metodo Folch (1957), uno dei più antichi e più utilizzati in matrici di diversa natura. Nato per essere usato in campioni con grasso molto ricco di fosfolipidi (come il grasso presente nel cervello), lavora a temperatura ambiente utilizzando una miscela di solventi composta da cloroformio:metanolo (2:1, v/v). Il metodo Acellerated Solvent Extraction (ASE) è stato utilizzato allo scopo di confrontare una metodica che lavora con alte temperature e pressioni (120°C e 20 MPa) con una miscela di solventi identica a quella usata nel metodo Folch (1957) (cloroformio:metanolo (2:1, v/v)). Grazie alle caratteristiche positive il metodo ASE, negli ultimi anni si è molto diffuso. Esso è caratterizzato da una maggiore velocità d’estrazione, un ridotto utilizzo di solventi e una minore laboriosità da parte degli operatori. Si pensa che le condizioni (temperatura e pressione) a cui lo strumento lavora possano provocare isomerizzazioni incrementando gli isomeri con configurazione trans/trans. L’ultimo è un metodo diretto che in seguito ad una fase preparativa (di liofilizzazione del campione fatta con lo scopo di rimuovere l’acqua) permette di ottenere gli esteri metilici degli acidi grassi (FAME) da poter analizzare in GCxGC. Questo metodo richiede un’esterificazione di tipo acido-basico. Il lavoro è stato suddiviso in quattro passaggi in modo da: identificare il potere di risoluzione e l’incidenza di picchi non identificati, lo studio delle fonti di variazione, test di Levene per determinare l’omoscedasticità o eteroscedasticità delle varianze e infine degli studi per valutare la correlazione tra metodi. L’incidenza di valori non identificati è legata al numero di picchi osservati, che dipendono dalla sensibilità del metodo e dal tessuto analizzato: fegato, grasso e muscolo (da 0.04 a 0.08, da 0.05 a 0.06 e da 0.05 a 0.12, rispettivamente). La maggiore incidenza di valori nulli è stata osservata nel fegato e in particolare negli acidi grassi a corta/media catena (C8:0 e C10:0) e nel muscolo negli acidi grassi C24:0, C20:1t1t unknown isomers e nei PUFAn3 (C20:3n3, C20:4n3, C22:5n6) sempre con il metodo Jenkins (2010). Tra tutte le fonti di variazione analizzate lo scopo, era considerare le due principali che sono state molto significative (P<0.001) e sono rappresentate dal tessuto, dal metodo e dalla loro interazione. L’effetto della dieta è stato significativo per gli acidi grassi facenti parte dell’integrazione (C18:0, C18:1cis9 e C18:2trans10,cis12). Dai test di Levene è emerso che le varianze sono eteroscedastiche, tranne la dieta che è risultata, omoscedastica. Il passo successivo è stato considerare una tecnica cromatografica alternativa al GCxGC che fosse più specifica per lo studio degli isomeri dei CLA (Chapter 7) Nonostante, il GCxGC abbia un maggiore potere di risoluzione non è in grado di identificare tutti gli isomeri dei CLA, ma solo i due principali (C18:2cis9,trans11 e C18:2trans10,cis12). Per questo motivo sono presenti altre tecniche cromatografiche che permettono un’identificazione più precisa come la cromatografia liquida su colonna d’argento (Ag+HPLC). Il lavoro è stato svolto in collaborazione con il Leibniz Insitute for Farm Animal Biology (Dummerstorf, Germany) e in particolare l’unità di Muscle and Biology Growth. L’obiettivo in questo caso era identificare il maggior numero d’isomeri dei CLA presenti nel campione. I tessuti analizzati erano: muscolo (Longissimus Thoracis), grasso sottocutaneo e fegato. Il grasso (estratto etereo, EE) è stato ottenuto con il metodo Folch (1957) cui è seguita poi un’esterificazione acido-basica. Le analisi statistiche sono state eseguite considerando gli effetti di dieta, tessuto e ripetizione sulla distribuzione degli isomeri. La prima cosa che differenzia questa metodica dal GCxGC è il numero di picchi identificati che sono molto più elevati (13 isomeri). Questo tipo di tecnica premette un’identificazione precisa di molti isomeri dei CLA con un unico problema nell’identificazione dei picchi che si trovano nella regione dei cis/cis (l’ultima a comparire nel cromatogramma), per la quale è difficile identificare l’isomero C18:2cis9,cis11. Dai risultati è emerso che il tessuto è molto significativo (P<0.001) e la distribuzione degli isomeri è tessuto dipendente, con una concentrazione più alta nel grasso. L’effetto della dieta, in particolare quella con integrazione 8.0 g di CLA al giorno, è risultato significativo per gli isomeri principali (C18:2cis9,trans11 e C18:2trans10,cis12). Nella seconda parte della tesi sono state prese in considerazione le fonti di variazione del profilo acidico, come razza, sesso, dieta, ordine di parto, età e tessuto (Chapter 3 e Chapter 4). Gli animali usati in queste prove appartengono a quattro razze ovine Venete: Alpagota, Brogna, Foza e Lamon che presentano diversa condizione di criticità; valutata in base al numero di capi allevati, di maschi arieti utilizzati per gli accoppiamenti e di allevamenti. Al fine di salvaguardare e recuperare queste razze, Veneto Agricoltura, su indicazione della Regione Veneto ha creato l’azienda sperimentale di Villiago, un importante centro di conservazione di queste razze, che mira a produrre nuovi giovani riproduttori (agnelle e montoni) per gli allevatori interessati e a organizzare azioni a sostegno dello sviluppo dell’allevamento. I dati riportati in questa tesi coinvolgono animali che sono stati allevati seguendo un piano di conservazione in-situ che prevedeva la collaborazione tra l’azienda sperimentale di Veneto Agricoltura “Villiago” e l’azienda sperimentale dell’Università di Padova “Lucio Toniolo”. In totale gli animali utilizzati sono stati 115 e sono stati allevati tra Dicembre 2010 e Luglio 2012. Macellati a età diverse, erano considerati appartenenti a tre categorie: 31 agnelli da latte, 36 agnelli leggeri, 24 agnelloni pesanti e 24 pecore. Le tre prove (agnello leggero, agnellone pesante, agnello da latte-pecora) erano caratterizzate da diete diverse. Nella prima prova “agnello leggero” le diete utilizzate erano: pascolo (PAS), fieno e concentrati con integrazione CLA rumino protetti (rpCLA, 8.0 g/d/animale) (CLA+) e dieta con fieno e concentrati senza integrazione di rpCLA (CLA-). Nelle prove “agnellone pesante” e “agnello da latte-pecora” le diete erano composte da concentrati con integrazione rpCLA (8.0 g/d/animale per gli agnelloni pesanti, 12 g/d/animale per le pecore e 4.0 g/d/animale per gli agnelli da latte) (CLA+) e concentrati senza integrazione rpCLA (CLA-). Dai risultati è emerso che diversi fattori possono influenzare il profilo acidico e in particolare dalla prima prova che la dieta può avere un ruolo molto importante (P<0.001). La presenza del pascolo è risultata significativa per molti acidi grassi a catena dispari e catena ramificata (forme iso e anteiso) che tendono a essere meno elevati al pascolo. Il motivo è che con la dieta è possibile alterate il PH ruminale e la flora microbica in esso presente modificando così anche gli acidi grassi che questa può produrre. Il ruolo del pascolo è importante anche per quanto riguarda gli acidi grassi a lunga catena (LC-PUFA). Il pascolo può aumentare gli Ω3 (rispettivamente, PAS=2.70, CLA-=1.46 e CLA+=1.54) riducendo gli Ω6 (rispettivamente, PAS =3.76, CLA-=4.41 e CLA+=4.75) e di conseguenze anche il rapporto Ω6/Ω3. Rispetto alla dieta a base di concentrati con integrazione (CLA+) il pascolo ha portato a un aumento dei CLA (rispettivamente, PAS=0.80, CLA-=0.56 e CLA+=0.71). La dieta si conferma significativa, anche nelle prove “agnellone pesante” e “agnello da latte-pecora” con il confronto tra le diete a base di concentrati (CLA- e CLA+). L’integrazione di rpCLA nella dieta è risultata significativa (P<0.001 e P<0.01) per gli acidi grassi considerati i suoi principali costituenti, per cui il C18:0 e il C18:2trans10,cis12. Gli agnelli da latte hanno presentato dei risultati diversi dagli altri animali, soprattutto in relazione ai CLA. Sono risultati significativi gli isomeri C18:2cis11,trans13 (P<0.01), C18:2cis11,cis13 (P<0.001) e C18:2cis9,cis11 (P<0.001). Il motivo di questa diversa distribuzione è stato attribuito al ruolo del latte di pecora, usato come alimento per gli agnelli e che può aver influito sulla composizione acidica dei loro tessuti. La razza in tutte le prove è stata un effetto che non ha portato a delle differenze significative dal punto di vista statistico, come anche il sesso e l’età di macellazione. Dai contrasti effettuati tra tessuti, è emerso, com’era stato osservato per i vitelloni, che la distribuzione degli acidi grassi è tessuto specifica e può cambiare secondo il tessuto analizzato. Differenze sono evidenti in tutti i tessuti ma in particolare tra i magri e quelli grassi. Il fegato in tutte e prove è stato il tessuto che presenta delle caratteristiche particolari perché caratterizzato da una maggiore quantità di acidi grassi polinsaturi (PUFA) e in particolare da una maggiore quantità di Ω3 e CLA. La diversa composizione di questo tessuto è legata alla composizione in termini di tipologia di lipidi che lo costituiscono e dai processi metabolici ai quali quest’organo partecipa che portano alla produzione, oppure all’assorbimento degli acidi grassi assunti con la dieta (Chapter 3 e Chapter 4). Sempre in animali appartenenti alle razze (Alpagota, Brogna, Foza e Lamon) sono state prese in esame le performance animali ed i vari fattori che le possono influenzare. E’ noto che l’utilizzo di rpCLA può provocare un incremento dell’efficienza alimentare favorendo l’aumento della massa magra con la successiva riduzione di quella grassa. Negli animali in lattazione (vacche, pecore e capre) è dimostrato che l’integrazione di rpCLA può provocare una riduzione nel contenuto di grasso presente nel latte. Nella mia tesi è stato considerato l’effetto della dieta sul latte di pecore (Chapter 5) al fine così di valutare quelle che vengono definite proprietà di qualità del latte espresse in termini di composizione chimica e proprietà di coagulazione (MCPs). Gli animali utilizzati in questa prova sono i medesimi utilizzati per lo studio dei profili acidici delle carni. Dal punto di vista della qualità della carne alla macellazione per cercare di definire se dieta, sesso e razza potessero influire sulle rese di macellazione (Chapter 6). Gli animali usati in questa prova sono i medesimi della prova vista in precedenza, ovvero la prova “agnello leggero”. I risultati ottenuti in questo lavoro sono dei risultati che comprendono solo parte del lavoro che invece comprenderà anche le prove “agnellone pesante” e “agnello-pecora”. Da questo lavoro quello che si vuole esprimere è che gli animali appartenenti a queste razze sono in grado di produrre delle carcasse che presentano caratteristiche ottimali e idonee al mercato alimentare. La caratteristica in più che queste presentano è che le modalità di allevamento, ben lontane da quelle di tipo industriale o più specializzato ci permettono di ottenere un prodotto allevato nel rispetto delle tradizioni e che può far parte di presidi ampiamente riconosciuti come l’Agnello Alpagoto noto per essere un presidio Slow Food.

Orientador: Fabio Augusto
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica
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Resumo: O objetivo do trabalho foi projetar e avaliar um modulador criogênico para cromatografia gasosa bidimensional abrangente (GC×GC). O modulador projetado foi baseado no modulador de quatro jatos (dois jatos quentes e dois jatos frios); N2 (g) resfriado em N2 (l) foi utilizado como fluido criogênico e N2 (g) aquecido foi utilizado como gás quente. O modulador foi instalado em um cromatógrafo a gás com detector por ionização em chama (GC-FID). O controle das válvulas solenóides e a digitalização do sinal analógico do FID foram realizados por software escrito em ambiente LabVIEW. Os resultados obtidos com o protótipo GC×GC-FID foram comparados com dados obtidos em um GC×GC-FID comercial. O desempenho de ambos os sistemas pode ser considerado equivalente, levando em consideração eficiência cromatográfica e repetibilidade. O protótipo GC×GC-FID foi empregado na análise de diversas amostras, em especial a fração volátil de polpa de abacaxi fresco e desidratado. Os voláteis foram extraídos por microextração em fase sólida através da extração dinâmica do headspace (DHS-SPME). A identificação dos compostos foi feita por cromatografia gasosa acoplada a espectrometria de massas (GC-MS) e avaliação dos índices de retenção. A identificação dos picos no cromatograma GC×GC foi feita através da comparação dos índices de retenção e dos perfis cromatográficos previamente obtidos por GC-MS. Como esperado, a análise por GC×GC-FID apresentou maior detectabilidade e poder de separação quando comparada com GC-FID. Por fim, alguns compostos não identificados por GC-MS foram identificados através de informação obtida pela estruturação cromatográfica da GC×GC.
Abstract: The aim of this project was to develop and evaluate a cryogenic modulator for comprehensive two-dimensional gas chromatography (GC×GC). The design of the modulator was based on a cryogenic quad jet modulator; N2 (g) cooled with N2 (l) was used as the cryogenic fluid and heated N2 (g) was used as the hot gas. The modulator was fitted into a gas chromatograph equipped with a flame ionization detector (GC-FID). The control of the solenoid valves and digitalization of the analogic FID signal were performed by software written in the LabVIEW platform. Results obtained with the GC×GC-FID prototype were compared with data from a commercial GC×GC-FID system. The performance of both systems was similar regarding chromatographic efficiency and repeatibility. The GC×GC-FID was employed for analysis of several samples, specially volatile organic compounds of fresh and dried pineapple pulp. The analytes were isolated by dynamic headspace solid phase microextraction (D-HSSPME); identification of the compounds was performed by conventional gas chromatography coupled to mass spectrometry (GC-MS) and evaluation of retention indexes. The identification of the peaks on the GC×GC chromatograms was carried out by comparison of retention indexes and chromatographic profiles previously obtained by GC-MS. As expected, the detectivity and separation power of the GC×GC-FID analysis was significantly better than that of GC-FID. Moreover, the identity of some compounds not identified by GC-MS was assigned after information obtained from GC×GC chromatographic structure.
Doutorado
Quimica Analitica
Doutor em Ciências

As determinações, em níveis de traços, de fármacos em fluidos biológicos e de multirresíduos (contaminantes) em amostras alimentícias são de extrema importância, pois geram valiosos dados para fins, respectivamente, de monitorização terapêutica (individualização do regime de dosagem) e controle de qualidade (segurança alimentar). A demanda por métodos analíticos de alta resolução e com baixos limites de quantificação, para análises de amostras complexas, tem impulsionado a química analítica para o desenvolvimento de soluções inovadoras, destacando-se aquelas voltadas ao desenvolvimento ou avaliação de novos sistemas analíticos. Neste contexto, na primeira etapa desta tese, o sistema automatizado de microextração em fase sólida no capilar de polipirrol (in-tube PPY SPME) acoplado à cromatografia líquida com detecção fluorimétrica foi desenvolvido (lab-made) para a determinação enantiosseletiva de fluoxetina e de seu metabólito norfluoxetina em amostras de plasma, para fins de monitorização terapêutica. Na segunda etapa, o método cromatografia gasosa bidimensional abrangente acoplada à espectrometria de massas com analisador quadrupolo (GC x GC / qMS) foi padronizado e validado para análise de multirresíduos de agrotóxicos em tomates frescos, para fins de controle de qualidade. Dentre os resultados obtidos podemos destacar: na primeira etapa o ganho de seletividade da fase extratora de polipirrol, em sistema miniaturizado e automatizado de preparo de amostra, hifenado à separação cromatográfica (LC) com detecção fluorimétrica; e na segunda etapa, o incremento da resolução cromatográfica e detectabilidade do sistema de cromatografia gasosa bidimensional com detecção espectrométrica com analisador quadrupolo. As análises de amostras de plasma de pacientes em terapia com fluoxetina e de amostras de tomates comerciais comprovaram a aplicabilidade dos métodos propostos, padronizados e validados, em níveis de concentrações que incluem o intervalo terapêutico preconizado para a fluoxetina em plasma e os limites máximos de resíduos de agrotóxicos estabelecidos para a cultura de tomate.
Determination of trace levels of drugs in biological fluids and multiresidue (contaminants) in food samples is extremely important because this generates valuable data for therapeutic drug monitoring (individualization of dosage regimen) and quality control (food safety), respectively. Because of the demand for analytical methods with high resolution and low limits of quantification for analysis of complex samples, analytical chemistry has stimulated the development of innovative approaches, especially those aimed at developing or evaluating new analytical systems. In this context, in the first stage of this thesis the automated solid-phase microextraction capillary polypyrrole (in-tube \"PPY SPME) coupled to liquid chromatography with fluorimetric detection was developed (lab-made) for enantioselective determination of fluoxetine and its metabolite norfluoxetine in plasma samples for therapeutic drug monitoring. In the second stage of this work, the GC x GC / qMS method was developed for multiresidues analysis of pesticides in fresh tomatoes for the purpose of quality control. In the first stage of the research the gain in terms of the selectivity of the polypyrrole extraction phase in hyphenated and automated system for sample preparation and chromatographic separation (LC) with fluorimetric detection is worthy of note. As for the second step, the highlight is the improvement in chromatographic resolution as well as in the detectability system of the system consisting of two-dimensional gas chromatography and spectrometric detection with quadrupole analyzer. Analyses of plasma samples from patients undergoing therapy with fluoxetine and of samples of commercially available tomatoes proved the applicability of the proposed methods, which were optimized and validated at concentrations levels that include the therapeutic range for the analyzed drugs in plasma and the maximum residue limits of pesticides for growing tomatoes.

Orientador: Fabio Augusto
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica
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Resumo: A Cromatografia Gasosa Bidimensional Abrangente (GCxGC) é uma técnica analítica capaz de separar e detectar centenas de picos, sendo aplicada a diferentes amostras petroquímicas, ambientais e biológicas. Esta técnica se diferencia da Cromatografia Gasosa (GC) pela utilização de duas colunas cromatográficas de composições e dimensões diferentes conectadas em série e de uma interface para a coleta das frações da primeira coluna e para a reinjeção na segunda em períodos regulares, denominada modulador. Além disso, por apresentar picos mais estreitos (wb 500ms), a frequência de aquisição utilizada é maior ( 100Hz), gerando arquivos que ultrapassam a capacidade de aplicativos comerciais. Assim, no desenvolvimento de sistemas próprios GcxGC, tem-se a necessidade de criar programas ou softwares para controle e tratamento de dados. Neste trabalho, foram desenvolvidos dois sistemas de controle distintos para sistemas GCxGC montados neste laboratório. O primeiro, denominado CROMATOGRAFIA, foi criado baseando-se no conceito de desenvolvimento de software da Fundação do Software Livre, ou seja, utilizou-se recursos de programação e de rede do sistema operacional Linux para realizar o controle e tratamento dos dados cromatográficos em um microcomputador e conversor A/D. Por outro lado, o segundo sistema, denominado LABVIEW, foi criado baseando-se no conceito de otimização de hardware, ou seja, utilizou-se um microcomputador e conversor A/D de médio desempenho em conjunto com uma linguagem de programação de alto nível específica para a automação de instrumentos. Ambos os sistemas de controle foram aplicados e validados em relação ao sinal monitorado, exatidão temporal e controle do modulador, verificando-se que o sistema LABVIEW apresentou resultados mais exatos para as análises qualitativa e quantitativa. Apesar de suas limitações iniciais, o sistema CROMATOGRAFIA com kernel soft real-time também foi aplicado com precisão e exatidão para controle dos sistemas GCxGC, salientando-se que seus módulos de tratamento e representação gráficas apresentaram melhor desempenho que aplicativos comerciais utilizados
Abstract: Comprehensive Two-dimensional Gas Chromatography (GCxGC) is an analytical technique used to separate hundreads of peaks and applied on different samples, like petrochemical products, enviromental and biological materials. This technique differs from convencional Gas Chromatography (GC) by use of two serial connected cappilary columns with different stationary phases and dimensions and a modulator, an interface that collects first column eluate fractions and reinjects them on second at regular time. Futhermore, data acquisition frequency must be higher ( 100Hz), because peak width acquired on GCxGC systems (wb 500ms), is narrow than conventional GC, resulting large data files to process with commercial softwares. On GCxGC system developments is necessary to create softwares for controlling and evaluating chromatographic data parameters. At this research two different control system were developed and applied to automate two home-made GCxGC systems. First control system, called CROMATOGRAFIA, was developed based on Free Software Foudation concept, i.e., it was developped on low level programming language and TCP/IP protocols within Linux O.S to control microcomputer and A/D conversor. On the other hand, second system, called LABVIEW, was developed on hardware optimize, i.e., it was developped with medium performance microcomputer and A/D conversor, using a commercial high-level visual programming language which is specific for automating instrument (NI Labview). Both control systems were checked performance by signal and real-time studies and applied for qualitative and quantitative analysis. LABVIEW system had better accuracy and precise results and, although its initial limits, CROMATOGRAFIA system with soft real-time kernel could be applied to accuracy and precise control GCxGC system with better 3D graphic representation and chromatographic data treatment than used commercial graphic softwares
Doutorado
Quimica Analitica
Doutor em Ciências

This work deals with the using and application of separation techniques for analysis of polymers degradation and polycyclic aromatic hydrocarbons. Thereby this work is separated to two special parts. In the first part, the degradation properties of synthetic biopolymers based on lactic acid, gylcolic acid and poly(ethyleneglycol) PLGA-PEG-PLGA and ITA-PLGA-PEG-PLGA-ITA (modified by itaconic acid) were studied. These copolymers (firstly their thermosensitive hydrogels) should be used for therapy of fractures in orthopedy (as adhesives). Therefore, the sol-gel and gel-sol phase diagrams were determinated for selected samples of copolymers. The samples forming gel at 37 C was used for other study. Polymer samples were depredated in phosphate buffer at 37°C. The degradation process of physical hydrogels was described by the decrease of molecular weight and the increase of concentration lactic acid and glycolic acid in phosphate buffer. The obtained results confirmed that the degradation of polymer modified by itaconic acid is faster process than no modified polymer and polymers with lower ratio PLGA/PEG degrade also faster than lower ration PLGA/PEG. The influence of pH it was also tested. The rate of degradation of polymers was follow pH 4,0

This diploma thesis deals with Bisphenol A, especially with its impact on the aquatic ecosystem. Information about its properties, production and current use are summarized here. Its harmful impact on the environment, especially on the aquatic ecosystem and the human body is characterized. Also the methods of its degradation within the aquatic environment have been described. A summary of the options of a determination of Bisphenol A in water samples is incorporated and the method of gas chromatography with mass spectrometry (GC/TOF MS) and comprehensive two-dimensional gas chromatography with mass spectrometry (GCxGC/TOF-MS) is compared in the experimental part. Analytical determination precedes the isolation of the analyte from the water samples by solid phase extraction (SPE) using SupelcleanTM ENVITM - 18 and derivatization using the silylation reagent, N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA). The result of this work is the analysis of a series of real samples from wastewater treatment plants Brno Modřice and Luhačovice by a two-dimensional gas chromatography with mass spectrometry (GCxGC/TOF-MS).

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
As árvores do gênero Copaifera (Leguminosae), conhecidas popularmente como copaibeiras, exsudam um óleo-resina extensamente utilizado na medicina popular e por indústrias farmacêuticas e de cosméticos, devido às suas atividades cicatrizante e anti-inflamatória. Quimicamente, esses óleos se caracterizam pela presença de hidrocarbonetos sesquiterpênicos, sesquiterpenos oxigenados e ácidos diterpênicos. No entanto, a composição química desses óleos-resina é variável e ainda não se tem conhecimento dos fatores que as determinam, embora vários fatores bióticos e abióticos sejam considerados fontes dessa variação. Essa variação dificulta a padronização da composição química desses óleos, comprometendo seu controle de qualidade e consequentemente a qualidade dos produtos a que darão origem, fato que tem causado um grande entrave à sua maior aplicação e comercialização. Essa variabilidade em sua composição química já é bastante conhecida e relatada na literatura, mas a maioria dos estudos realizados têm se restringido a caracterizar quimicamente o óleo-resina e poucos tem se preocupado em estudar as causas dessas variações. O objetivo deste trabalho foi padronizar a composição química dos óleos de copaíba por meio de técnicas de cromatografia em fase gasosa acoplada à detectores de ionização de chama (CG-DIC) e espectrometria de massas (CG-EM) e cromatografia em fase gasosa bidimensional abrangente (CGXCG); analisar estatisticamente a influência de fatores abióticos como sazonalidade, tipo de solo e diâmetro à altura do peito (DAP), além da infestação por cupins, sobre a composição química desses óleos. Além disso, foram comparados 5 métodos de esterificação dos ácidos diterpênicos presentes nos óleos de copaíba envolvendo catálise ácida, que utilizam BF3/MeOH, H2SO4/MeOH e HCl/MeOH, levando-se em consideração suas eficiências e frequências analíticas, além do consumo e toxicidade dos reagentes utilizados, relação custo benefício e, principalmente, a possibilidade de alteração/degradação da estrutura dos constituintes quando aplicados em óleos de copaíba. Para isso foram obtidos óleos de copaíba de 3 coletas: em novembro de 2004 e novembro de 2005 (épocas consideradas secas) e em maio de 2005 (época considerada chuvosa). No total, 43 amostras de óleo-resina de copaíba foram coletadas na Reserva Ducke (Manaus-AM), de 33 espécimes diferentes, que possuíam diferentes DAP s e se encontravam em diferentes tipos de solo. As análises por CG-DIC e CG-EM permitiram a identificação de 35 constituintes: sendo 22 hidrocarbonetos sesquiterpênicos, 9 sesquiterpenos oxigenados e 4 ácidos diterpênicos. Enquanto a análise por CGxCG permitiu a identificação de outros 13 sesquiterpenos, além de 7 monoterpenos, inéditos em óleos-resina de copaíba. O β-cariofileno e seu óxido foram os constituintes majoritários em 29 e 11 amostras, respectivamente. As análises hierárquica por agrupamento (HCA) e de componentes principais (PCA) evidenciaram a existência de dois grupos distintos com diferentes perfis cromatográficos, em que foi comprovada apenas a influência do tipo de solo, sobre a composição química desses óleos. Outros fatores analisados como sazonalidade, DAP e infestação por cupins, não tiveram influência sobre a composição química dos óleos-resina de copaíba. Quanto aos métodos de esterificação, as análises das 5 metodologias testadas, apesar de terem sido reprodutíveis, não se mostraram eficientes, ao passo que não permitiram a identificação dos constituintes formados e levaram à formação de artefatos.

Ce mémoire de doctorat rassemble les travaux réalisés dans le cadre d’une étude analytique approfondie et systématique des substances à odeur boisée. Ces substances définissent une famille d’ingrédients unanimement appréciés pour leurs notes puissantes et recherchés par les parfumeurs pour composer les accords parmi les plus emblématiques de la profession. Les matières premières inscrites au programme de cette étude sont les essences de Gaïac, d’Araucaria, de Patchouli, de Cyprès Jaune d’Alaska, de Vétiver et d’Agarwood, ainsi que certains de leurs produits dérivés. La plupart de ces matières premières sont couramment utilisées par l’industrie du parfum. Leur spécificité provient de leur composition chimique très largement dominée par les dérivés sesquiterpéniques, conférant ainsi à ces extraits une complexité moléculaire indéniable. Les principaux objectifs de cette thèse de doctorat ont consisté à améliorer la connaissance de ces matières premières en termes de composition chimique, à déterminer autant que possible les composés odorants contribuant à leur odeur, et enfin d’explorer d’éventuelles voies de valorisation pour ces mêmes ingrédients à l’échelle industrielle.La méthodologie analytique développée tout au long de l’étude a nécessité l’utilisation combinée d’un large éventail de techniques chromatographiques, spectrométriques et spectroscopiques pour parvenir à caractériser ces matières premières au plus près et à réaliser l’isolement de leurs constituants inconnus. Ainsi, ces travaux s’articulent autour de quatre outils centraux : la chromatographie gazeuse bidimensionnelle intégrale couplée à la spectrométrie de masse (GC × GC–MS), la chromatographie gazeuse couplée à l’olfactométrie (GC–O), la chromatographie gazeuse semi-préparative (GC prép.) et la résonance magnétique nucléaire (RMN). Près de 190 composés ont été isolés et caractérisés dont 107 sont décrits pour la première fois comme constituants d’extraits naturels
This Ph.D. dissertation sums up the work carried out as part of a thorough and systematic analytical study of woody-scented substances. These substances define a family of ingredients unanimously appreciated for their powerful notes, and sought by perfumers to compose some of the most emblematic accords in perfumery. The raw materials included in the frame of this study are the oils of Guaiac, Araucaria, Patchouli, Alaska Yellow Cypress, Vetiver, and Agarwood, as well as some of their by-products. Most of these raw materials are ingredients commonly used by the perfume industry. Their chemical specificity comes from their composition largely dominated by sesquiterpene derivatives, giving these extracts an undeniable molecular complexity. The main objectives of this Ph.D. thesis were to improve the knowledge of these raw materials in terms of chemical composition, to determine as much as possible the odorous compounds contributing to their odor, and finally to explore possible ways of valorization for these products on an industrial scale.The analytical methodology developed throughout this study required the combined use of a wide range of chromatographic, spectrometric, and spectroscopic techniques in order to characterize these raw materials as precisely as possible and achieve the isolation of their unknown constituents. Thus, our work revolved around four central tools: comprehensive two-dimensional gas chromatography-mass spectrometry (GC × GC-MS), gas chromatography-olfactometry (GC–O), preparative capillary-gas chromatography (pc-GC) and nuclear magnetic resonance (NMR). Almost 190 compounds have been isolated and characterized, 107 of which are described for the first time as constituents of natural extracts

This work is based on the current issue of increasing concentrations of pharmaceutical residues in various components of the environment. These new environmental contaminants continuously enter the environment. The most affected environmental component is the aquatic environment. This study is focused on the development and optimization of reliable analytical method, which can determine selected drugs in the aquatic environment (waste waters and surface waters) qualitatively and quantitatively. The target compounds were selected mainly from the group of non-steroidal anti-inflammatory drugs (NSAID): salicylic acid, acetylsalicylic acid, clofibric acid, ibuprofen, acetaminophen, caffeine, naproxen, mefenamic acid, ketoprofen and diclofenac. NSAIDs are one of the most used drugs in Czech Republic. For the final analysis of this study was used the comprehensive two-dimensional gas chromatography with mass spectrometric detection Time-of-Flight (GCxGC-TOF MS). It is a very sensitive and reliable analytical method for trace and ultra-trace analysis. Simultaneously, solid phase extraction (SPE) and derivatization are optimized in this work. Optimized analytical method including SPE, derivatization with MSTFA (N-methyl-N-(trimethylsilyl) trifluoroacetamide) and final analysis by GCxGC-TOF MS were applied successfully for the analysis of real samples. Samples of waste water were collected from the waste water treatment plant in Brno – Modřice and samples of surface water were collected from two river streams Svratka and Svitava in Moravian region. The range of concentrations of selected drug residues varied from one to tens of g/L in wastewater and from tens to hundreds of ng/L in surface waters. The degree of wastewater and surface waters contamination by drug residues is assessed in conclusions of the work. Simultaneously, spontaneous degradation of selected drugs in water and removal efficiency of the wastewater treatment plant is assessed in conclusions of the work. The developed method is usable for monitoring and environmental analysis of water ecosystems. It is usable not only for selected drugs, but even for other organic compounds with similar properties.

Os hidrocarbonetos policíclicos aromáticos nitrados (nitro-HPAs) são potencialmente genotóxicos, por isso existe um grande interesse na determinação analítica desses compostos no material particulado no ar. Os nitro-PAHs são emitidos diretamente para a atmosfera pela combustão incompleta ou podem ser formados in situ a partir dos HPAs precursores. Neste trabalho, são apresentados os resultados obtidos no desenvolvimento de um método de análise de nitro-HPAs (1-nitronaftaleno, 2-nitronaftaleno, 1-nitropireno, 2-nitropireno, 2-nitrofluoranteno, 3-nitrofluoranteno, 1,3-dinitropireno, 1,6- dinitropireno, 2-nitrofluoreno, 6-nitrocriseno, 7-nitrobenzo(a)antraceno, 9- nitroantraceno e 9-nitrofenantreno) por GCxGC. A separação destes compostos por GCxGC pode simplificar a etapa de pré-tratamento do extrato orgânico oriundo do material particulado no ar. A GCxGC promove a separação em duas colunas com mecanismos de separação diferentes, conectadas em série, o que resulta em uma capacidade de separação maior. O sistema GCxGC empregado neste trabalho possui um injetor com programação de temperatura (PTV), um modulador criogênico de dois jatos de CO2 líquido (Dual Jet) e um detector por ionização em chama (FID). A separação foi feita empregando a fase estacionária apolar 5% fenil polisilfenilene siloxano na primeira coluna (30 m / 0,25 mm / 0,25 µm) e a semipolar 14% cianopropilfenil polisiloxano na segunda coluna (1 m / 0,10 mm / 0,10 µm). Diferentes condições cromatográficas foram testadas, como: programação de temperatura no PTV, de 40-90ºC até 350ºC, com taxa de aquecimento de 2,2 e 14,5ºC/s; programação de temperatura no forno, de 40-90ºC até 300ºC, com taxa de aquecimento de 2, 2,5 e 3ºC/min; períodos de modulação de 3, 4 e 5s. O volume injetado foi 2µL, no modo splitless; a vazão do gás de arraste (He) foi constante e igual a 1 mL/min e o detector FID foi operado em 120 Hz. As larguras de pico medidas em 10% da altura variaram de 167 a 350ms para 1- nitronaftaleno e 6-nitrocriseno, respectivamente, e a capacidade de pico para o conjunto de colunas de separação usado foi estimada em cerca de 9800. Parâmetros de validação, tais como os limites de detecção (200 a 500 ng/mL) e de quantificação (650 a 1600 ng/mL) para alguns nitro-HPAs foram determinados. Para a quantificação, foram testados os métodos por padronização interna e externa, sendo que, através dos resultados deste trabalho, pode-se mostrar que a determinação de nitro-HPAs por GCxGC com o modulador empregado é possível se for feita através do método de padronização interna. O sistema GCxGC empregado causou dificuldades operacionais, principalmente na conexão das colunas e na modulação. Perdas de massa de analito foram detectadas nas análises por GCxGC, muito provavelmente causadas por uma conexão parcial das colunas. Isso poderia ser evitado se a conexão fosse feita pela fusão das colunas a um conector de vidro adequado a essa finalidade. Picos modulados divididos foram observados com frequência, o que comprometeu a precisão e exatidão dos resultados. A divisão dos picos ocorreu de modo aleatório dificultando a identificação da causa mais provável para a sua ocorrência. Cromatogramas GCxGC de extratos orgânicos não fracionados de particulados no ar, oriundos de queima direta de palha de cana-de-açúcar, de região afetada pela queima de cana-de-açúcar e de área urbana da cidade de São Paulo, mostraram a potencialidade do método
Nitro-polycyclic aromatic hydrocarbons (nitro-PAHs) are potentially genotoxic, so there is a great interest in the analytical determination of these compounds in airborne particulate matter. The nitro-PAHs are emitted directly into the atmosphere by incomplete combustion or may be formed in situ from PAHs. In this work, results obtained from the development of an analytical method for nitro-PAHs by GCxGC are presented. The nitro-PAHs studied were: 1- nitronaphthalene, 2-nitronaphthalene, 1-nitropyrene, 2-nitropyrene, 2- nitrofluoranthene, 3-nitrofluoranthene, 1,3-dinitropyrene, 1,6-dinitropyrene, 2- nitrofluorene, 6-nitrochrysene, 7-nitrobenzo(a)anthracene, 9-nitroanthracene and 9- nitrophenanthrene. The separation of these compounds by GCxGC should simplify the pretreatment step of the airborne particulate organic extract. In GCxGC two capillary columns of different mechanisms of separation are coupled by an interface called modulator, which increase significantly the chromatographic resolution. The GCxGC system used in this work is equipped with a PTV injector, a dual-stage double jets CO2 cryogenic modulator and a flame ionization detector (FID). The separation was carried out using a nonpolar stationary phase 5% methyl phenyl polysiloxane in the first column (30 m / 0.25 mm / 0.25 µm) and a semipolar 14% cyanopropyl phenyl dimethyl polysiloxane in the second column. Different chromatographic conditions were tested: the PTV was programmed from 40-90ºC to 350ºC at 2.2 and 14.5ºC/s; the oven was programmed from 40-90ºC to 350ºC at 2.0, 2.5 and 3.0ºC/min; the modulation periods tested were 3, 4 and 5s. The injected volume was 2µL in splitless mode, the carrier gas (He) flow rate of 1 mL/min was constant and the FID detector was operated at 120 Hz. Validation parameters, such as the limit of detection (200 - 500 ng/mL) and quantitation (650 - 1600 ng/mL) were determined. For quantification, the internal and external standardization methods were tested, and through the results of this study, it can be shown that the determination of nitro-PAHs by GCxGC with the modulator used is possible if made by internal standardization method. The GCxGC system presented operational difficulties, particularly in the column connection and in the modulation zone. A partial connection of the columns resulted in mass loss of the analytes. The connection of the columns could be more effective if it was made by fusing the columns to a glass connector suitable for this purpose. Divided modulated peaks were frequently observed, which compromised the precision and accuracy of the results. The division of the peaks occurred randomly hindering the identification of the most likely cause for its occurrence. GCxGC chromatograms corresponding to unfractionated organic extracts of airborne particulate material from the sugarcane straw burning, from an area impacted by sugar cane burning emissions and from an urban area of São Paulo City showed the capability of the method.

碩士
國立中央大學
化學研究所
98
Abstract Comprehensive two-dimensional chromatography (GCxGC) opened a new era in improving separation resolution for very complex samples. It uses two columns of different polarity to provide orthogonal separation on a 2-D surface, which considerably increases the peak capacity and hence resolution compared to conventional 1-D chromatography. A typical commercial GCxGC system often uses a cryogenic modulator whose cost is high, and the consumption of cryogen creates additional financial burden. In contrast, many studies focused on developing valve-based types of modulators, which are simple in design and construction. Moreover, cryogen is not required. In our study we used a Deans’ switch to construct the modulator for GCxGC. When combined with a self-constructed thermal desorption unit, this system was able to analyze atmospheric chlorofluorocarbons (CFCs) at pptv level. The volatile sample peaks eluted from the first column (DB-1 60 m) were sliced into sample pulses by the Deans’ switch and sent to the second column (PLOT 0.2 m) for the second dimensional separation. For optimal GCxGC separation of CFCs the modulation pulse and period were set at 0.2 s and 3 s, respectively.By varying the sample size of a pressurized air sample, the linearity of the method was better than 0.990. The precision at atmospheric concentrations was better than 4.74% (RSD). The Deans’ switch-GCxGC method was further tested by continuous measuring outdoor ambient air, and compared the atmospheric variability of CCl2F2 (CFC-12) and CClF2CCl2F (CFC-113) with that from the corresponding 1D GC results. The variability of CFC-12 and CFC-113 denoted by RSD was 3.85% and 3.49%, respectively, for GCxGC versus 2.63% and 3.34%, respectively, for 1D GC. The comparable results suggest that our Deans’ switch-GCxGC method is sufficiently robust to analyze atmospheric CFCs.

Technology has become increasingly more prevalent in all aspects of society since the age of the computer. The United States Military has successfully integrated the powerful processing capabilities of computers to increase the proficiency and lethality of its Soldiers, Sailors, Marines, and Airmen. However, this increased lethality comes at risk due to the inherent vulnerabilities of computer systems to spyware, malware, and counterfeit components. Inspired by the ability of canines to seek out and find electronic devices, this research sought methods to characterize components by their “scent” using precise analytical tools. Using these tools, this thesis sought to develop and utilize non-invasive methods to show proof-of-concept for electronic device classification by volatile compounds unique to different types of components. The findings of this research proved that electronic components that vary by age, origin, type, or manufacturer emit different volatile compounds available for detection using modern two-dimensional gas chromatography and solid-phase microextraction technologies. If developed further, the methods used in this research have the potential for application in the United States Department of Defense to ensure that all electronic components installed in their systems are authentic, come from a trusted source, and can be relied upon in even the most stressful operating conditions.

Environmental petroleum hydrocarbon (PHC) monitoring is a major challenge. Analytical methods must be robust; operate with minimal user intervention; be suitable for remote field operation; and furnish analytical data that allows the different mechanisms of PHC environmental fate to be investigated. PHC are amenable to analysis by comprehensive two-dimensional gas chromatography (GCxGC). However, conventional GCxGC instrumentation relies on bulky thermal modulation systems. Thus alternative approaches based on fluidic modulation were investigated to determine their suitability for environmental PHC monitoring. First, a dynamic flow model, which maps carrier gas pressure and flow rate through the first-dimension separation column, the modulator sample loop, and the seconddimension column(s) in a fluidic modulation GCxGC system is described. The dynamic flow model assists design of a pneumatic modulation ensemble and leads to rapid determination of pneumatic conditions, timing parameters, and the dimensions of the separation columns and connecting tubing used to construct the GCxGC system. Three significant innovations are introduced, that were all uncovered by using the dynamic flow model, viz. i) a “symmetric flow path” modulator improved baseline stability, ii) appropriate selection of flow restrictors in the first dimension column assembly provides a generally more stable and robust system, and iii) these restrictors increase the modulation period flexibility of the GCxGC system. Next, a model was developed that permitted a systematic investigation of peak shape in fluidic modulation. In the case of a non-focusing modulator for comprehensive two-dimensional gas chromatography, the systematic distortions induced when the modulator loads the second-dimension column give rise to a characteristic peak shape. Depending on the operating conditions this systematic distortion can be the dominant component of the second-dimension elution profiles. Understanding the factors that cause different peak shape observations provides a rugged approach to method development. It is shown that low flow ratio can lead to significant peak skewing and increasing the flow ratio reduces the magnitude of peak skewing. Validation of the peak shape model is made by comparison with experimental data. Finally GCxGC methodology was developed and applied to analysis of PHC contaminated soil. GCxGC results met or exceeded, the standards set by regulators and environmental scientists. Fluidic modulation approaches provided excellent sensitivity and permitted detailed monitoring of key PHC transport and degradation pathways, including evaporation, dissolution, and biodegradation.

碩士
國立臺灣師範大學
化學系
98
The goal of this research is to investigate three different configurations of two dimensional gas chromatography (2D-GC) when using two of micro electromechanical system (MEMS) technology fabricated -column chips coated with DB-1 and DB-210 as stationary phases. Taking advantage of dissimilar retentions for VOCs of the two stationary phases and 2D-GC, we resolve the problem of insufficient separation using single, short -column chip. Commercial modulators for 2D-GC are cryogenic or valve-based. However, cryogenic modulators need to have cylinders of liquid nitrogen or carbon dioxide. It violates the unique purpose of portability for μGC. Hence, μGC should use only cryogenic free modulators. 　　The three modulator systems for 2D-GC investigated in current study are Heart cutting GC × GC system, Deans switch GC - GC system, and Stop-flow GC - GC system. The Heart cutting GC × GC system and Stop-flow GC - GC system can separate eleven VOCs of similar boiling points within five minutes. Although the Heart cutting GC × GC system causes the loss of sample mass due to splitting, it achieves better separations. The Deans switch GC - GC system links to two detectors so that it increases the efficiency of experiment. Comparing with the same VOCs, this system only takes a half of time to finish the separation of these. Stop-flow GC - GC system is the lightest in hardware requirements, but it is also the most time consuming among three modulator systems.

碩士
國立臺灣大學
電機工程學研究所
99
Three works are included in this thesis including 1) an algorithm for Comprehensive two-dimensional gas chromatography mass spectrometry alignment, 2) 3Omics: a web based systems biology visualization tool for integrating human transcriptomic, proteomic and metabolomic data, and 3) HMO: a tool for understanding the human metabolome. A novel peak alignment algorithm, 2DGCMS-aligner, has been developed for two-dimensional gas chromatography time-of-flight mass spectrometry (GCxGC/TOF-MS) data. 2DGCMS-aligner uses the netCDF data generated from the instrument as input directly. It detects blobs, clusters of pixels that are brighter or darker than their surround in a chromatogram, of each GCxGC/TOF-MS raw data to generate blob tables instead of peak tables to perform alignment. 2DGCMS-aligner correlates the blobs with Euclidean distance of the first- and second retention times in the blob tables and the mass spectra with Pearson’s correlation coefficient. This alignment algorithm in 2DGCMS-aligner can be applied to GCxGC-MS data generated by either consistent or inconsistent instrument environment to adjust retention time shifts along both chromatographic dimensions caused by uncontrollable fluctuations in temperature and pressure, matrix effects and stationary phase degradation. 2DGCMS-aligner also includes an option to correct baseline on raw data directly. The performance of 2DGCMS-aligner peak alignment algorithm was compared and demonstrated with three existing alignment methods on the two sets of GCxGC-MS data sets acquired in different experiment conditions and a mixture of standard metabolites. 3Omics: a web based systems biology visualization tool for integrating human transcriptomic, proteomic and metabolomic data was developed to visualize and rapidly integrate multiple inter- or intra-transcriptomic, proteomic, and metabolomic human data. A biochemical cascade is generated through consolidation of transcript, protein, and metabolite data and implements via the application of five commonly used analyses of correlation network, co-expression, phenotyping, KEGG pathway enrichment, and GO enrichment. 3Omics incorporates the advantages and operations of existing software into a single platform, therefore simplifying the data analysis procedure and enabling the user to perform a one-click integrated analysis for free. Visualization and analysis results are downloadable for further user customization and analysis. The 3Omics software can be freely accessed at http://cmdd.csie.ntu.edu.tw/~3omics. Last part of this thesis work is the construction of Human Metabolome Ontology (HMO). Final step in current metabolomics studies involves assessment and biological interpretation of metabolome. It often requires tedious manual collections of literature or linking information scattered in Gene Ontology, BRENDA, KEGG Brite, KEGG Pathway, Human Metabolome Database, OMIM and so on. We developed the HMO to facilitate integration of biological functions, and chemical classification of metabolome and comprehensive understanding of metabolome and its target interactions as the common language and knowledge framework allowing further computational analysis. HMO consists of three independent ontologies: biological functions, chemical taxonomies and metabolome targets. It provides a comprehensive metabolome centered resource that enables the sharing and reuse of the know-ledge across domains of ontologies.

碩士
國立中央大學
化學研究所
96
The continual anthropogenic chlorofluorocarbons (CFCs) are known to destroy stratospheric ozone because of their long lifetimes and chemical stability. The advent of the Montreal Protocol caused total phase-out of CFCs in 1996. The consumption of the replacements, hydrochlorofluorocarbons (HCFCs) and hydrofluorocarbons (HFCs), have been increasing rapidly. Although they have limitted or no ozone depleting potentials (ODPs), they are very potent green house gases. In this study, a gas chromatographic (GC) method was developed to simultaneously measure CFCs, HCFCs and HFCs in the atmosphere. The system involved an automated preconcentration unit using chemical adsorbents combined with two dimensional GC equipped with electronic captured detector (ECD) for measuring CFCs. For HCFCs and HFCs the oxygen-doping ECD method was employed to enhance the sensitivity. The system was tested on campus and at the Taishi EPA monitoring station. Limited emissions of CF2ClCFCl2 and CCl4 were found, which can serve as a useful quality assurance tool to validate measurements. Other more variable species, i.g., CF2Cl2, CF2ClBr and CHCl3, were detected revealing the existence of sources. For the doping system deployed in Taishi station, strong emissions of CH2Cl2 with concentration excess of 8 ppb relative to the baseline condition was found. The datasets were further examined by Gaussian distribution, which is a novel approach to reveal variability and sources of halocarbons.

碩士
國立中央大學
化學學系
104
Comprehensive two-dimensional gas chromatography (GC×GC) is the preferred choice of analysis for complex volatile organic compounds (VOCs). When coupled with time-of-flight mass (ToF) spectrometry, termed GC×GC-ToF, it becomes a powerful technique to analyze fuels, perfumes, aromas, environmental samples, etc. with complex chemical compositions. Compared to conventional one-dimensional GC, GC×GC greatly enhances peak capacity via the use of two columns of different phases and lengths for orthogonal separation. Modulation plays a central role in GC×GC performance. The high cost in ownership and operation of a commercial GC×GC system equipped with a cryogenic modulation motivated the development of cryogen-free modulation. In this study, a valve-based modulator based on the Deans switch served as an alternative to the commercial counterpart without the use of cryogen. The switching of an auxiliary gas stream of a Deans switch that cuts peaks from the column of first dimension (1D) into fine slices to the short column of secondary dimension (2D) created the effect of modulation. Because of the low concentrations of VOCs in ambient air, usually at only sub-ppbv levels, an air sample would require substantial preconcentration before GC analysis. As a result, a self-built preconcentrator was connected to a GC×GC system, which was equipped with two columns of DB-1 (60 m×0.32 mm i.d.×1 μm d.f.) as the 1D column and Rtx-502.2 (2 m×0.32 mm i.d.×1.8 μm d.f.) as the 2D column to display the orthogonality of non-polarity vs. mid-polarity. Flame ionization detection (FID) was adopted by exploiting its high acquisition rates and reliability. Instead of using the commercial GC×GC software packages, a general-purpose software, Surfer®8, was used to plot GC×GC the results. The analytical precision of 7% as the RSD was achieved by repeated analysis of the PAMS standard gas mixture at sub-ppbv level. Ambient samples collected in a long highway tunnel by canisters and sorption tubes were analyzed for system validation. Furthermore, as the trial studies, PM2.5 aerosol samples collected on filter papers were attempted by thermal desorption of the filter paper. Compound identification was made with GC-MS by analyzing parallel samples to reveal the chemical identities of the major constituents of both the air and PM samples. While all the VOCs found in the canisters were non-polar hydrocarbons due to the lack of photochemistry in the tunnel, selected oxygenated VOCs (o-VOCs) were found in the aerosol sample owing to the extended oxidation process in the atmosphere.

碩士
國立中央大學
化學研究所
95
Alkylphenol polyeyhoxylates (APEOs), one of the most important classes of nonionic surfactants, were introduced in the 1940s. Because of their great soaking, cleaning and emulsifying abilities, they are widely used in the household detergents, textile industries, dyeing industries and other commercial applications in Taiwan. APEOs in aquatic environment are biodegraded into alkylphenols (APs) and relative metabolites. Indeed, APs have been estimated that they are persistence organic pollutions which are not easily biodegraded by microorganisms and demonstrated as endocrine disruptors which can cause estrogenic effects in organisms. This effect has raised increasing concern about their impact on wildlife and human health. Consequently, it is necessary to develop analytical methods for routinely determining the levels of alkylphenolic compounds in the Taiwanese environment and observe the biodegradation of 4-nonylphenol isomers. The first part of this study is using steam distillation extraction and derivatization methods coupled with gas chromatography/mass spectrometry (GC/MS) to determine the levels of alkylphenol compounds in fish. The silylating agent used was BSTFA+1% TMCS and the optimal reaction time and temperature were 30 min and 80°C. Under these conditions, the recovery of OP and NP was above 66.8% and 67.4%, respectively. RSD ranged from 2.1 to 10.4%. The concentration of 4-t-OP and 4-NPs ranged from 13-202.9 ng/g and from 9.2-529.7 ng/g, respectively. The second part of this study is deterimining the alkylphenols in commercial milk and breast milk. The method involves extracting a sample by liquid-liquid extraction and solid-phase extraction and then clean up by anhydrous sodium sulfate and aluminum oxide. Alkylphenols were identified and quantitated by gas chromatography /mass spectrometry (GC/MS) in selected ion monitoring (SIM) mode. Various solution compositions were evaluated as rising solution. The limit of quantitation (LOQ) were less than 0.2 ng/g for 4-t-OP and 1.5 ng/g for 4-NP in 20 g breast milk. In 20 breast milk samples, 4-t-OP was detected in 8 samples and concentration ranged from 0.4 to 1.14 ng/g, while 4-NPs was detected in 19 samples and concentration ranged from 1.74 to 11.58 ng/g. In 7 commercial milk samples, 4-t-OP was detected in 1 samples and concentration was 0.14 ng/g, while 4-NPs was detected in all testing samples and concentration ranged from 2.93 to 8.84 ng/g. The results of the recovery and reproducibility for the SPE method indicated that it’s better to use methanol:water=1:1 (v/v) solution than ethanol:water=1:1 (v/v) as rinsing solution. The spiked recovery of 4-t-OP and 4-NPs were above 77.5% while RSD was below 9% which indicated this method is reliable and sensitive for determining traces of alkylphenols residues in milk. Finally, because of different biodegradation rates, estrogenic effect and bioaccumulation potentials of each 4-nonylphenol isomer, only a few 4-nonylphenol isomers have higher estrogenic effects. So we used 100 m GC column and two-dimensional gas chromatography (GC x GC) combined with quadrupole mass spectrometer to separate 4-nonylphenol isomers more further in fish and breast milk samples. The results indicated that the more branched isomers of 4-nonylphenol isomers which may be more resistant to biodegradation than the less branched isomers in organisms. Furthermore, we analyzed five technical 4-nonylphenol standards from different vendors by GC x GC-MS system. Different patterns were observed from these standards. Hence, the GC x GC-MS system can be used as a rapid screening tool to access nonylphenol pollution associated with chemical migration and source apportionment studies.