Soil-aquifer treatment (SAT) has been proposed as a method for reusing treated municipal wastewater. SAT is characterized by alternating cycles of aerobic and anaerobic conditions in the subsurface, in response to alternating cycles of flooding and drainage of a surface impoundment. It is not yet known how these alternating redox conditions affect the removal of potentially harmful endocrine-disrupting compounds (EDCs) from treated effluent. The overall objective of my doctoral research is to determine the fate of EDCs in alternating aerobic/anoxic/anaerobic conditions under simulated SAT conditions. To assess the fate of EDCs in simulated SAT conditions, I first had to develop appropriate analytical methods. Prior researchers have developed sophisticated analytical methods for measuring low concentrations of EDCs in water. However, it is not inherently clear which of these methods is preferable for analysis of any particular set of environmental samples. Therefore, in order to compare the analytical methods, solid-phase extraction (SPE) and solid-phase micro-extraction (SPME) were compared for the analysis of two EDCs, bisphenol-A (BPA) and 17B-estradiol (E2), in water samples of water. Following extraction by SPE or SPME, the target EDCs were derivatized (silylated) and then analyzed by gas chromatography (GC) with mass spectrometry (MS). Also, the performance of two candidate derivatization agents, N,O-bis-(trimethylsiyl) trifluoroacetamide (BSTFA), N-methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA), was compared. SPME is more convenient, is less labor-intensive, and allows for analysis of smaller sample volumes, but it is expensive because fibers need frequent replacement, and the range of linearity was limited. SPE has a lower material cost and allows for the analysis of a broader range of concentrations, but it is more labor-intensive and large sample volumes may be required. Therefore, the selection of which method is "best" depends upon the constraints (time, money, sample volume, acceptable detection limit) associated with any particular set of samples. The two derivatization agents performed equally when used in conjunction with SPE, but MSFTA yielded higher peak areas for headspace (on-fiber) derivatization during SPME. To investigate how alternating redox conditions of SAT may affect the removal of harmful EDCs, a simulated SAT systems were constructed in 4-L reactors with 500 g of sediment (collected from a wetland) and 3 L of treated effluent from a municipal wastewater treatment plant; then BPA and E2 were spiked into reactors, two common EDCs often found in treated wastewater. Redox conditions in the mesocosms were controlled by switching the air between air (to induce aerobic conditions) and nitrogen (to induce anaerobic conditions); the length of the anoxic/anaerobic cycles was varied to determine how this affects biodegradation of the target EDCs. The mesocosm environment was supplemented with either nitrate or sulfate to serve as potential electron acceptors during the anoxic/anaerobic cycles. In addition to monitoring the concentrations of the target EDCs in the mesocosms over time, I also monitored the concentration of dissolved oxygen in the water; the redox potential; the concentrations of nitrate, nitrite, and sulfate; and the concentration of bacteria in the water (estimated via flow cytometry). BPA was biodegraded only during aerobic cycles, but E2 was biodegraded during both aerobic and anoxic/anaerobic cycles. Whenever the redox conditions in the system were switched, there was a temporary drop in the bacterial population, followed by a recovery of the population. When redox conditions were switched from anoxic/anaerobic to aerobic, biodegradation of the target EDCs commenced after a lag period during which no biodegradation was observed. The lag time for biodegradation in the aerobic cycle was longer when the anaerobic cycles were longer in duration. More biodegradation of E2 was observed under anoxic conditions than under anaerobic conditions. SPE and SPME methods that included derivatization agent are useful method for detection and quantification of EDCs in water. I concluded that SAT is a viable technology to produce potable water from treated WWTP effluent, but the optimal length of flooding and drying cycles of SAT required removing the targeted contaminants during infiltration through the vadose zone.

L'aroma és un factor de qualitat de la majoria dels aliments, ja que en depèn, en gran mesura, que el consumidor accepti o rebutgi el producte. En el vi i en les begudes alcohòliques esdevé, però, un dels factors més importants, ja que aquests productes no es consumeixen pel seu valor nutritiu, sinó pel plaer que en proporciona la degustació.L'anàlisi de l'aroma del vi no és un problema senzill de resoldre si es consideren els centenars de compostos volàtils que poden intervenir en la seva configuració aromàtica i els baixos nivells de concentració en què es troben molts d'ells. El treball d'investigació realitzat en aquesta tesi ha consistit en l'estudi i la caracterització de l'aroma del vi mitjançant l'aplicació de dues tècniques molt importants en aquest àmbit: la cromatografia de gasos amb detecció olfactimètrica (GCO) i el nas electrònic.La GCO és una tècnica imprescindible en l'estudi de l'aroma que, utilitzant el nas humà com a detector cromatogràfic, permet identificar, d'entre tots els compostos volàtils de la mostra, aquells que presenten olor i que, per tant, són susceptibles de participar en l'aroma. El treball més rellevant dut a terme amb aquesta tècnica ha estat el desenvolupament d'un mètode, per a caracteritzar l'aroma del vi, que utilitza la microextracció en fase sòlida (SPME) com a tècnica de pretractament de la mostra i que permet l'avaluació de la intensitat aromàtica dels diferents compostos mitjançant una nova modalitat de la tècnica de dilució AEDA (Aroma Extract Dilution Analysis).Els instruments coneguts com a nassos electrònics es van desenvolupar amb la finalitat d'imitar el nas humà en el control de qualitat que, des de temps immemorials, s'ha dut a terme mitjançant l'anàlisi sensorial. En l'anàlisi de begudes alcohòliques, però, els nassos electrònics han estat poc utilitzats degut als problemes de saturació que causa l'elevat contingut d'etanol als sensors de gasos que conformen la majoria d'aquests instruments. En aquesta tesi s'ha estudiat l'aplicació d'un nas electrònic basat en l'espectrometria de masses al control de qualitat del vi i de les begudes alcohòliques, concretament brandies i aiguardents de canya de sucre, ja que, a priori, l'etanol no suposa una limitació per a l'ús d'aquest instrument. La major part dels estudis realitzats s'han dirigit a l'avaluació del potencial d'aquesta tècnica en el control de la qualitat del vi. Amb aquest objectiu s'han dut a terme amb èxit diferenciacions de vins segons diversos paràmetres enològics, com són l'origen, la varietat o el temps d'envelliment. Per altra banda, s'ha fet un seguiment de la maduració del raïm i del procés d'elaboració del vi per tal d'avaluar la capacitat d'aquesta tècnica en el control d'aquests dos processos. Una altra aplicació estudiada ha estat la determinació de compostos amb un impacte sensorial important, com els defectes aromàtics no desitjats, mitjançant el calibratge multivariant. En aquesta línia s'han desenvolupat dos mètodes per a la determinació del 2,4,6-tricloroanisole, un compost que es troba en alguns vins a causa de contaminacions externes i que és el principal responsable del defecte aromàtic conegut com a "gust de suro". Per últim, s'ha estudiat també l'aplicació de l'instrument al control de qualitat dels aiguardents de canya de sucre amb el desenvolupament d'un mètode per a determinar el temps d'envelliment dels aiguardents en bótes de fusta de roure, així com també s'ha estudiat la correlació entre les dades obtingudes en l'anàlisi dels aiguardents amb el nas electrònic i les intensitats de diferents descriptors aromàtics avaluades per un panel de tastadors en una anàlisi sensorial de les mostres. / Aroma is an important factor in quality control and quality assurance of foods, but in wines and in alcoholic beverages, this factor is possibly the most important. Wine aroma analysis is not an easy task to perform because aroma is composed of complex mixtures of hundreds of volatile compounds with different sensory and chemical properties and, moreover, the concentration levels of most of them are very low.The research work carried out in this thesis lay in the study and the characterisation of wine aroma by two very important techniques in this field: gas chromatography olfactometry (GCO) and electronic nose. GCO is a very useful technique in the study of wine aroma that makes it possible to identify, among all the volatile compounds, the odorous volatile compounds of the sample by using the human nose as chromatographic detector. The most important study carried out in this thesis by GCO was the development of a method for the characterisation of wine aroma that uses the solid-phase microextraction technique (SPME) as a sample pre-treatment technique. This method evaluates the aromatic intensity of the odorous compounds by a new approach of the aroma extract dilution analysis (AEDA) technique.Quality control of food has been carried out by sensory analysis since immemorial time. However, sensory analysis by a panel of experts is a costly process for industries because it requires trained people who can work for only relatively short periods of time. The purpose of the development of electronic noses was to achieve an instrument that could mimic the human sense of smell and provide rapid sensory information (e.g., differences and similarities among samples, presence of aromatic defects, etc.). Food analysis is probably the field in which the greatest number of applications using electronic noses have been developed. However, in wine and alcoholic beverage research, few studies have been performed with this type of instrumentation because of the problems that ethanol causes in the gas sensors which compose most of the electronic noses. In this thesis, the application of a mass spectromety (MS) based electronic nose to wine analysis has been studied because, a priori, ethanol does not interfere in the analysis by this instrument. Some applications have also been carried out with alcoholic beverages as sugar cane spirits and brandies. The purpose of these studies was to evaluate the potential application of this technique in the quality control of this kind of samples. In wine analysis, this technique was successfully applied to differentiate and classify wines according to their origin, variety and ageing. It was also evaluated its capacity in the monitoring of the aroma profile evolution during grape ripening and alcoholic fermentation in order to study the potential use of this technique in the control of these processes. Other application studied was the determination of compounds with an important sensory impact, such as the off-flavours, by multivariate calibration. Two methods were developed to determine 2,4,6-trichloroanisole which is the main responsible of the aromatic defect known as "cork taint". In the quality control of sugar cane spirits, a method to determine the ageing time of these samples in oak wood barrels was developed. It was also studied the correlation between the electronic nose data and the intensity of several odour attributes evaluated by a panel of experts in a sensory analysis of the spirits.

The feasibility of using solid phase micro-extraction (SPME) as an in vivo sampling tool for analysis of trace environmental contaminants in fish exposed to municipal wastewater effluents (MWWEs) was validated using controlled laboratory and field experiments. SPME was compared with traditional extraction techniques， including solid phase extraction (SPE) in water and solid-liquid extraction (SLE) in fish tissues to assess relative efficiencies. All three techniques were used to quantify the presence of eight compounds of interest in fish exposed to MWWEs in the laboratory, as well as in wild and field caged fish upstream and downstream of three wastewater treatment plants in the Grand River watershed. Atrazine, carbamazepine, naproxen, diclofenac, gemfibrozil, bisphenol A, fluoxetine and ibuprofen were selected as target compounds due to their diverse chemical characteristics and frequent detection in surface waters and sediments around the world. The distribution coefficients between various sample matrices (water, fish) and extraction phases (SPME fibers) were compared, as were extraction profiles and bioconcentration factors of target analytes in muscle of fish exposed to MWWEs under laboratory conditions, during field caging studies, or collected (wild) from the Grand River. Poly(dimethylsiloxane) (PDMS) medical grade tubing was utilized as the SPME extraction phase, which when kinetically calibrated, were effective at extracting and quantifying the target analytes from both water and fish tissue relative to traditional techniques. Caged and in wild fish exposed to MWWEs from all three municipal treatment plants bio-accumulated detectable levels of several of the target chemicals. All target analytes (except for fluoxetine) were identified in the MWWEs and exposed fish by SPME at low concentrations (ng/L). The presence and concentration of the targeted analytes in both water and wild fish living in the Grand River watershed varied with season and proximity to the wastewater outfalls. Results demonstrate that properly applied SPME can detect and quantify selected contaminants in fish tissues, surface water, and wastewater effluents. In vivo SPME allows for non-lethal sampling of fish, which creates the opportunity for monitoring contaminant exposure in receiving environments influenced by MWWEs or non-point-source runoff while minimizing the impact on the organisms.

This thesis describes the development and application of solid phase microextraction (SPME) as a sample preparation technique for in vivo determination of pharmaceutical residues in fish tissue and wastewater. The occurrence, distribution and fate of pharmaceuticals in the environment are a subject of concern across the globe due to the impact they may have on human life and aquatic organisms. To address this challenge from an analytical perspective, a simplified and reliable analytical methodology is required to investigate and determine the concentration (bioconcentration factors) of trace pharmaceutical residue in fish tissue and environmental water samples (exposure). An improved SPME method, coupled with liquid chromatography with tandem mass spectrometry has been developed and applied to both controlled laboratory and field-caged fish exposed to wastewater effluent for quantitative determination of pharmaceutical residue in fish specific tissue. A new SPME configuration based on C18 thin film (blade) was developed and optimized to improve SPME sensitivity for in vivo determinations of trace pharmaceuticals in live fish. The C18 thin film extraction phase successfully quantified bioconcentrated fluoxetine, venlafaxine, sertraline, paroxetine, and carbamazapine in the dorsal-epaxial muscle of living fish at concentrations ranging from 1.7 to 259 ng/g. The reproducibility of the method in spiked fish muscle was 9-18% RSD with limits of detection and quantification ranging from 0.08 - 0.21 ng/g and 0.09 - 0.64 ng/g (respectively) for the analytes examined. Fish were sampled by in vivo SPME for 30 min to detect pharmaceutical uptake and bioconcentration, with experimental extracts analyzed using liquid chromatography coupled with tandem mass spectrometry. In addition, a simplified analytical methodology based on SPME was developed and optimized for determination and bioconcentration factor of different classes of pharmaceuticals residues in fish bile. The reproducibility of the method in spiked fish Rainbow Trout bile was 3-7% RSD with limits of detection (LOD) ranging from 0.3 – 1.4 ng/mL for the analytes examined. The field application of SPME sampling was further demonstrated in Fathead Minnow (Pimephales promelas), a small-bodied fish caged upstream and downstream of a local wastewater treatment plant where fluoxetine, atorvastatin, and sertraline were detected in fish bile at the downstream location. Also, a simple automated analytical method using high throughput robotic system was developed for the simultaneous extraction of pharmaceutical compounds detected in surface waters. The proposed method successfully determined concentrations of carbamazepine, fluoxetine, sertraline, and paroxetine in treated effluent at concentrations ranging from 240 - 3820 ng/L with a method detection limit of 2-13 ng/L, and a relative standard deviation of less than 16%. Application of the method was demonstrated using wastewater from pilot-scale municipal treatment plants and environmental water samples from wastewater-dominated reaches of the Grand River (Waterloo, ON). Finally, 4 and 8-d laboratory exposures were carried out with Rainbow Trout exposed to wastewater effluent collected from pilot scale at Burlington, ON. Additionally, wild fish, White Sucker (Catostomus commersonii) were collected and sampled from Waterloo and Kitchener downstreams containing local municipal effluent. Bioconcentration factors of the selected compounds were determined in both fish muscle and bile samples. The results show that anti-depressant drugs such fluoxetine, sertraline and paroxetine were uptake in the fish muscle and fish bile for both laboratory and field exposure. In summary, exposure of fish to micro-pollutants such as pharmaceuticals may be monitored through the analysis of bile, particularly at low concentration exposure of pharmaceuticals, where the sensitivity of analytical method may be challenged. SPME is a promising simple analytical tool which can potentially be used for monitoring of pharmaceuticals in fish tissue and wastewater.

The ability to locate human remains, specifically in a forensic setting, is crucial to investigations. Research in the past two decades has identified volatile organic compounds (VOCs) as the source of the decomposition odor. The study examined the headspace (area directly above) of decomposing remains, Sigma-Aldrich (St. Louis, MO) Pseudo Corpse Scents (formulations one and two), cadaveric blood, and decompositional fluid for target VOCs to which human remains detection (HRD) dogs could indicate. These samples were tested using solid-phase microextraction (SPME) and a gas chromatograph-mass spectrometer (GC/MS) for the exact odor profile and compared to literature about VOCs present in decomposition. The author hypothesized that a series of seven target chemical compounds (carbon disulfide, hexanal, nonanal, dimethyl sulfide, dimethyl disulfide, styrene, and benzoic acid methyl ester) would be present when the headspace of all samples tested. Ideally, a synthetic compound that will better mimic human decomposition odor profile can be created to aid in the training of HRD dogs. There are some disadvantages to using dogs in the field, and the lack of standardization when training HRD dogs is a major one. By examining VOC profiles of different sources of decomposition, a core set of VOCs of human decomposition may be identified to aid in the standardization of training. Of the eight target compounds chosen from the literature, only two were found in any samples tested for this experiment, hexanal and nonanal found in pig heart, deer liver, as well as human muscle and epidermis. Acetic acid was identified in every sample with the exception of the control. Also, in accordance with the literature, putrescine and cadaverine were not found in any of the samples.

Submitted by Ana Hilda Fonseca (anahilda@ufba.br) on 2016-09-06T15:39:12Z No. of bitstreams: 1 Tese Adalberto Menezes Filho 26.03.2010.pdf: 4529406 bytes, checksum: 23a6a27845c477df8cc06bee3255777f (MD5) / Approved for entry into archive by Vanessa Reis (vanessa.jamile@ufba.br) on 2016-09-08T10:44:55Z (GMT) No. of bitstreams: 1 Tese Adalberto Menezes Filho 26.03.2010.pdf: 4529406 bytes, checksum: 23a6a27845c477df8cc06bee3255777f (MD5) / Made available in DSpace on 2016-09-08T10:44:55Z (GMT). No. of bitstreams: 1 Tese Adalberto Menezes Filho 26.03.2010.pdf: 4529406 bytes, checksum: 23a6a27845c477df8cc06bee3255777f (MD5) / CNPq / Foram desenvolvidas, validadas e aplicadas duas metodologias analíticas por SPME e análise por GC-MS e HPLC-UV-Vis, para determinar resíduos de agrotóxicos em manga. 14 compostos foram analisados por GC-MS (clofentezina, carbofuran, diazinon, parationa, malationa, fentiona, tiabendazol, imazalil, bifentrina, permetrina, procloraz, piraclostrobina, difenoconazol, azoxistrobina) e 10 por HPLC-UV-Vis (tiabendazol, carbofuran, azoxistrobina, procloraz, fentiona, clofentezina, permetrina, abamectina, carbosulfan e bifentrina). Diferentes parâmetros que influenciam na eficiência da extração foram avaliados (Tipo de fibra, modo de extração, temperatura e tempo de extração e dessorção, velocidade de agitação e força iônica). Os melhores resultados foram obtidos com fibra de PA e DI a 50°C por 30 min, com agitação a 250 rpm e dessorção por 5 min a 280°C no GC- MS e no modo estático por 15 min na interface SPME-HPLC. Na validação foram avaliados o efeito da matriz, a linearidade das curvas analíticas, LOD, LOQ, precisão e exatidão. O método por SPME-GC-MS apresentou linearidade entre 3,3 e 1665,0 µg kg-1, LOD entre 1,0 e 3,3 µg kg-1 e LOQ entre 3,3 e 33,3 µg kg-1. O método por SPME-HPLC- UV-Vis apresentou linearidade entre 2,0 e 250,0 µg kg-1, LOD entre 0,6 e 3,3 µg kg-1 e LOQ entre 2,0 e 10,0 µg kg-1. Nos dois métodos foram obtidos CV menores que 20%. Os métodos foram aplicados na análise de amostras coletadas em Salvador-BA e Aracaju-SE. Nas amostras de Salvador foram detectados resíduos de sete compostos e nas de Aracaju resíduos de cinco compostos. Entretanto, as concentrações estavam abaixo dos valores estabelecidos pela Legislação Brasileira. / Were developed, validated and applied two analytical methodologies by SPME and GC-MS and HPLC-UV-Vis analysis to determine pesticide residues in mango. 14 compounds were analyzed by GC-MS (clofentezine, carbofuran, diazinon, methyl parathion, malathion, fenthion, thiabendazole, imazalil, bifenthrin, permethrin, prochloraz, pyraclostrobin, difenoconazole, azoxystrobin) and 10 for HPLC-UV-Vis (thiabendazole, carbofuran, azoxystrobin, prochloraz, fenthion, clofentezine, permethrin, abamectin, bifenthrin and carbosulfan). Different parameters influencing the extraction efficiency were evaluated (fiber type, extraction mode, temperature, extraction and desorption times, stirring velocities and ionic strength. The best results were obtained using PA fiber and DI mode at 50°C form 30 min, along with stirring at 250rpm and desorption for 5 min at 280°C in the GC-MS and estatic mode for 15 min in the SPME-HPLC interface. For validation, we assessed the matrix effect, the linearity of calibration curves, LOD, LOQ, precision and accuracy. The method for SPME-GC-MS showed linearity between 3.3 and 1665.0 mg kg- 1, LOD between 1.0 and 3.3 µg kg-1 and LOQ between 3.3 and 33.3 µg kg-1. The method for SPME-HPLC-UV-Vis showed linearity between 2.0 and 250.0 µg kg-1, LOD between 0.6 and 3.3 µg kg-1 and LOQ between 2.0 and 10.0 µg kg-1. In both methods were obtained CV below 20%. The methods were applied in the analysis of samples collected in Salvador- BA and Aracaju-SE. In samples from Salvador seven compounds residues were detected and in samples from Aracaju five compounds residues were detected. However, the concentrations were below the values established by Brazilian legislation

Submitted by Paulo Mesquita (prrmesquita@gmail.com) on 2016-10-18T17:10:54Z No. of bitstreams: 1 Tese Doutorado Mesquita_FINAL.pdf: 3135632 bytes, checksum: dd198da88e048f82bac99316af7e483b (MD5) / Approved for entry into archive by Uillis de Assis Santos (uillis.assis@ufba.br) on 2016-10-18T17:18:46Z (GMT) No. of bitstreams: 1 Tese Doutorado Mesquita_FINAL.pdf: 3135632 bytes, checksum: dd198da88e048f82bac99316af7e483b (MD5) / Made available in DSpace on 2016-10-18T17:18:46Z (GMT). No. of bitstreams: 1 Tese Doutorado Mesquita_FINAL.pdf: 3135632 bytes, checksum: dd198da88e048f82bac99316af7e483b (MD5) / FAPESB e CNPq. / O comércio de frutas tropicais, principalmente das regionais e exóticas, apresenta-se como uma grande oportunidade para produtores e outros profissionais brasileiros. No entanto, a sua comercialização tem crescido aquém das expectativas devido, em certa parte, à necessidade de maior fomento a pesquisas relacionadas a estas espécies frutíferas e seus possíveis produtos derivados. Entre as diversas abordagens que estão sendo estudadas na área agrícola, se destacam a caracterização dos metabólitos presentes nas diferentes espécies de plantas de interesse econômico, o entendimento do papel biológico dos mesmos e possíveis alterações neste metaboloma gerados por ataques de diferentes tipos de patógenos. Este trabalho teve como objetivo caracterizar os metabólitos presentes na pitangueira (Eugenia uniflora L.) e diferentes tipos de maracujazeiros (Passiflora spp.), e avaliar possíveis mudanças no perfil destes compostos. Foram determinados os perfis de compostos orgânicos voláteis (COVs), extraídos de folhas de E. uniflora de plantas que apresentavam diferentes biotipos de cor dos seus frutos (laranja, vermelho ou roxo), coletadas em diferentes regiões da Bahia. Os compostos foram extraídos e identificados através da técnica HS-SPME/GC-MS e o perfil das amostras foi avaliado através de técnicas de análise multivariada (PCA e HCA). Através das técnicas aplicadas foi possível identificar 33 compostos nas plantas com cada biotipo de cor de fruto e discriminar entre os três grupos de amostras com base neste perfil de metabólitos. Também foram extraídos e caracterizados, utilizando a técnica HS-SPME/GC-MS, os perfis de COVs de cinco espécies do gênero Passiflora: P. edulis, P. cincinnata, P. maliformis, P. gibertii e P. setacea. Em seguida avaliou-se alterações no perfil de COVs destas espécies após infecção pelo Cowpea aphid-borne mosaic virus (CABMV). Blocos de 4 plantas sadias e inoculadas com o CABMV foram mantidas em casa de vegetação e fitotron, sob condições controladas. Também foram avaliadas mudanças no perfil de compostos em P. cincinnata até o 28º dia após a inoculação com o CABMV. Foram identificados 43 COVs nas cinco espécies de Passiflora e a utilização das técnicas de análise multivariada (PCA, PLS-DA e HCA) permitiu discriminar entre as amostras sadias e infectadas pelo CABMV nas diferentes espécies, através do seu perfil de metabólitos. Além disso, foi possível identificar potenciais biomarcadores de infecção ao CABMV comuns e específicos para cada uma das cinco espécies. Ao avaliar a infecção de plantas da espécie P. cincinnata em diferentes períodos, se verificou que a partir de 3 dias de inoculação já é possível distinguir claramente o perfil das amostras infectadas comparado ao das amostras sadias. A abordagem metabolômica utilizada neste trabalho possibilitou identificar diferentes tipos de biomarcadores, tanto de espécies de Passiflora quanto prováveis variedades de E. uniflora para estudos de quimiotaxonomia, além de metabólitos sinalizadores de infecção pelo CABMV em Passiflora spp. / The trading of regional and exotic tropical fruits can be presented as a great opportunity for producers and other Brazilian professionals. However, their marketing has grown short of expectations due to some part to the need for higher encouragement of research related to these fruit species and its possible derivatives. Among the various approaches being studied in agriculture, stand characterization of metabolites present in different plant species of economic interest, understanding of the biological role of these and possible changes in the metabolome of different types generated by pathogen attacks. This study aimed to characterize the metabolites present in the species Eugenia uniflora L. and species of Passiflora, and evaluate possible changes in the profile of these compounds. It was determined the profiles of VOCs extracted from E. uniflora leaves of plants showing different color biotypes of its fruits (orange, red or purple), collected in different regions of Bahia. The compounds were extracted by HS-SPME/GC-MS technique and the profile of the samples was evaluated using multivariate analysis (PCA and HCA). Through the techniques applied were identified 33 compounds in plants with each fruit color biotype and discriminate between the three groups of samples based on this metabolite profile. Also were characterized using the HS-SPME/GC-MS technique, the VOC profiles of five species of the genus Passiflora: P. edulis, P. cincinnata, P. maliformis, P. gibertii and P. setacea. Then we evaluated changes in VOC profile of these species after infection by Cowpea aphid-borne mosaic virus (CABMV). Blocks of four healthy and inoculated plants with CABMV were kept in a greenhouse and phytotron, under controlled conditions. They evaluated changes in compounds profile in P. cincinnata until the 28th day after inoculation with CABMV. Were identified 43 VOCs in five species of Passiflora spp. and use of multivariate analysis (PCA, PLS-DA and HCA) allowed the discrimination between healthy and infected with CABMV samples in different species, through its metabolite profile. Moreover, it was possible to identify potential biomarkers of infection CABMV the common and specific to each of the five species. When evaluating infection of plants of the species P. cincinnata at different times, it was found that from 3 days of inoculation is already possible to distinguish the profile of the infected sample compared to the healthy samples. The metabolomic approach used in this study enabled us to identify different types of biomarkers, both species of Passiflora spp. as likely varieties of E. uniflora for chemotaxonomy studies, besides flags metabolites of infection by CABMV in Passiflora spp.

183 f. / Submitted by Ana Hilda Fonseca (anahilda@ufba.br) on 2013-08-26T14:09:17Z No. of bitstreams: 1 Tese_Alain_Gaujac.pdf: 13017399 bytes, checksum: 3307e5b9dd6143b6038b6951b60e2334 (MD5) / Approved for entry into archive by Ana Hilda Fonseca(anahilda@ufba.br) on 2013-08-26T14:11:56Z (GMT) No. of bitstreams: 1 Tese_Alain_Gaujac.pdf: 13017399 bytes, checksum: 3307e5b9dd6143b6038b6951b60e2334 (MD5) / Made available in DSpace on 2013-08-26T14:11:56Z (GMT). No. of bitstreams: 1 Tese_Alain_Gaujac.pdf: 13017399 bytes, checksum: 3307e5b9dd6143b6038b6951b60e2334 (MD5) Previous issue date: 2013 / CNPq / N,N-dimetiltriptamina ou DMT é um alcaloide indólico com acentuada ação psicoativa, presente em bebidas vegetais de origem indígena, como o vinho da jurema e a ayahuasca. Esses preparos vegetais são consumidos em rituais religiosos sincréticos criados no Brasil no início do século 20, hoje dispersos e em contínua expansão por todo o mundo. As bebidas são também utilizadas como droga de abuso, no contexto recreativo, principalmente, graças à fácil disponibilidade das fontes vegetais utilizadas nos seus preparos, oferecida pelo comércio virtual. Neste trabalho, foi proposto um estudo de caracterização do DMT, isolado e purificado a partir das cascas da M. tenuiflora (jurema-preta), envolvendo metodologias instrumentais de análise como ressonância magnética nuclear de hidrogênio e carbono (RMN de 1H e 13C), espectroscopia na região do infravermelho (IR) e espectrometria de massas (MS). O nível de pureza do padrão foi determinado por absorção no ultravioleta (UV), utilizando um padrão de triptamina. Na execução dos trabalhos iniciais, percebeu-se a possibilidade de existência de polimorfismo para o DMT, fato comprovado através de técnicas de análise calorimétrica (DSC) e difração de Raios X. A partir do padrão analítico desenvolvido, foram propostos dois métodos para a determinação de DMT em matrizes vegetais e nas bebidas utilizadas, como sacramento, em rituais religiosos de origem brasileira, ambos otimizados por meio de técnicas multivariadas. O método MSPD/GC-MS desenvolvido para a quantificação de DMT nas cascas de M. tenuiflora foi devidamente validado, apresentando excelentes figuras de mérito. Apresentou boa linearidade (r = 0,9962) e repetibilidade (CV < 7,4%), com limite de detecção de 0,12 mg g-1. Foram analisadas 24 amostras de cascas, nas quais se verificou a presença de DMT, em níveis de concentração entre 1,26 e 9,35 mg g-1. O segundo método, descreve os procedimentos para a determinação de DMT nas bebidas rituais, por HS-SPME/GC- MS. Foi realizado amplo estudo de validação. Boa precisão (CV < 8,6%) e excelente exatidão, com recuperações entre 71–109%. Os limites de detecção e quantificação foram 0,78 e 9,5 mg L-1, respectivamente, além de boa linearidade (1,56–300 mg L-1, r2 = 0,9975). A análise do efeito na resposta analítica, causado por pequenas variações nos parâmetros otimizados, revelou excelente robustez. O método validado foi aplicado na análise de amostras reais de ayahuasca (7) e vinho da jurema (5). Todas as amostras foram diluídas para análise. Verificou-se grande variabilidade entre as concentrações de DMT nas bebidas. Nas amostras de vinho da jurema, os níveis de DMT estiveram entre 0,1 e 1,81 g L-1. Nas amostras de ayahuasca, concentrações entre 0,17 e 1,14 g L-1. A análise de outras cinco amostras de vinho da jurema, preparadas em laboratório por diferentes procedimentos, e diluídas para os ensaios, revelou que o aquecimento não tem grande significância na quantidade de DMT extraído da planta, sendo mais importantes o baixo pH do líquido extrativo e a presença de etanol. / Salvador

A Aniba canelilla (H.B.K.) Mez é uma planta aromática proveniente da região amazônica cujo óleo essencial apresenta como componentes majoritários o 1-nitro-2-feniletano e o metileugenol. Apesar das atividades antifúngicas e anti-inflamatórias cientificamente comprovadas e do uso popular do óleo para o tratamento de dermatites, acnes e feridas, não existe até o momento um estudo que verse sobre a quantificação desses compostos na pele. O objetivo deste trabalho foi a validação de um método bioanalítico otimizado por microextração em fase sólida no modo headspace em cromatógrafo gasoso com detector de ionização de chama (HS-SPME-GC-FID) para a determinação do 1-nitro-2-feniletano e metileugenol a partir do óleo essencial em diferentes amostras de estudo de permeação cutânea in vitro. Uma metodologia foi desenvolvida e validada por HS-SPME-GC-FID. A faixa da curva de calibração foi de 2,08 - 207,87 μg.mL-1 para o 1-nitro-2-feniletano e de 0,40-40,41 μg.mL-1 para o metileugenol. A presença de matriz e as características intrínsecas da metodologia de HS-SPME requereram uma transformação da curva de calibração. A transformação logarítmica (Log10) foi então aplicada aos dados e os resultados apresentaram homocedasticidade, resíduos dispersos, coeficiente de determinação (r²> 0,99) e recuperação adequados. Estudos de permeação cutânea foram realizados em células de Franz com diferentes quantidades (20, 100 e 200 μL) de óleo essencial de A. canelilla para avaliar o perfil de permeação e retenção em pele de orelha suína e fluido receptor. A análise das amostras nas condições validadas mostrou uma grande permeação e retenção dos compostos na seguinte ordem: fluido receptor >> derme >> epiderme >> estrato córneo. Verificou-se um aumento progressivo e dependência na retenção com base na quantidade aplicada no compartimento doador e grande retenção principalmente no fluido receptor e derme, resultado este compatível com as características físico-químicas de Log P, afinidade ao ambiente hidrofílico e lipofílico, tamanho e peso molecular do 1-nitro-2-feniletano e metileugenol. Em conclusão, o método proposto por HS-SPME-GC-FID para quantificar os compostos majoritários do óleo essencial de A. canelilla em amostras de pele de orelha suína e fluido receptor foi seletivo, preciso, exato e adequado, e pode ser utilizado em análises futuras. / Aniba canelilla (H.B.K.) Mez is an aromatic plant from the Amazon region whose essential oil has 1-nitro-2-phenylethane and methyleugenol as major compounds. Despite of the scientifically proved antifungal and anti-inflammatory activities and the popular use of oil for the treatment of dermatitis, acnes and wounds, there is no study up to date about the quantification of these compounds in skin samples. The aim of this study was the validation of an optimized bioanalytical method by solid phase microextraction on headspace mode in gas chromatograph with flame ionization detector (HS-SPME-GC-FID) for the determination of 1-nitro-2-phenylethane and methyleugenol from the essential oil in different samples of in vitro cutaneous permeation study. A methodology was developed and validated by HS-SPME-GC-FID. The ranges of calibration curves were 2.08 - 207.87 μg.mL-1 for 1-nitro-2-phenylethane and 0.40 - 40.41 μg.mL-1 for methyleugenol. The presence of matrix and the intrinsic characteristics of the HS-SPME methodology required a transformation to the calibration curves. The logarithmic transformation (Log10) was then applied to the data and the results showed homoscedasticity, dispersed residues, and adequate coefficient of determination (r²> 0.99) and recovery. Skin permeation studies were performed on Franz cells with different amounts (20, 100 and 200 μL) of A. canelilla essential oil to evaluate the skin permeation and retention profile in porcine ear skin and receptor fluid. The analysis of the samples under the validated conditions showed a high permeation and retention of the major compounds in the following order: receptor fluid >> dermis >> epidermis >> stratum corneum. A progressive increase and retention dependence were observed based on the amount applied in the donor compartment, and large retention mainly on the receptor fluid and dermis was observed, in accordance with the physicochemical characteristics of Log P, affinity to the hydrophilic and lipophilic environment, size and molecular weight of 1-nitro-2-phenylethane and methyleugenol. In conclusion, the method proposed by HS-SPME-GC-FID to quantify the major compounds of A. canelilla essential oil in porcine ear skin and receptor fluid samples was selective, accurate, precise and adequate, and can be used in future analyzes.

Submitted by Maurílio Figueiredo (maurilioafigueiredo@yahoo.com.br) on 2014-07-02T18:25:21Z No. of bitstreams: 2 license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) DISSERTAÇÃO_EstudoComparativoMétodos.pdf: 2292877 bytes, checksum: 835505b7bbaee9d0eecceaf24630beaa (MD5) / Approved for entry into archive by Gracilene Carvalho (gracilene@sisbin.ufop.br) on 2014-07-17T18:38:52Z (GMT) No. of bitstreams: 2 license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) DISSERTAÇÃO_EstudoComparativoMétodos.pdf: 2292877 bytes, checksum: 835505b7bbaee9d0eecceaf24630beaa (MD5) / Made available in DSpace on 2014-07-17T18:38:52Z (GMT). No. of bitstreams: 2 license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) DISSERTAÇÃO_EstudoComparativoMétodos.pdf: 2292877 bytes, checksum: 835505b7bbaee9d0eecceaf24630beaa (MD5) Previous issue date: 2012 / Produzidos desde a década de 40, os organofosforados foram os primeiros a substituírem os organoclorados, aos quais os insetos já apresentavam resistência. Dentre estes o clorpirifós apresenta-se como um inseticida altamente tóxico. Devido à sua elevada lipossolubilidade é absorvido pelo organismo humano, especialmente pela pele. Os organofosforadossão conhecidos por induzirem ou agravarem certos problemas de saúde; assim é importante seu monitoramento. Para a determinação de clorpirifós em amostras aquosas geralmente utiliza-se etapas de extração/pré-concentração antes de se efetuar a quantificação. Neste trabalho, foramavaliadas as técnicasextração líquido-líquido com partição a baixa temperatura (LLE-LTP) e microextração em fase sólida com headspace (HS-SPME) e as técnicas de cromatografia gasosa com detecção por captura de elétrons (GC-ECD) e cromatografia gasosa acoplada à espectrometria de massas (GC-MS) para definição do método analítico mais sensível, utilizando planejamentos experimentais multivariados para a otimização dos métodos de preparo de amostra. Após a otimização, o método HS-SPME/GC-ECD foi validado para o monitoramento de clorpirifós em cursos d’água superficiais próximos às principais plantações de batata na zona rural de Ouro Branco/MG. Apresença de agrotóxicos, como o clorpirifós nesses cursos d’água, foi relacionada em pesquisas anteriores aos altos índices dos casos de hipertensão e aborto da população da zona rural. A definição dos pontos de coleta foi feita após visitas de campo e levantamento de informações obtidas junto aos órgãos e pessoas competentes. O planejamento fatorial completo apontou as variáveis tempo (1)e temperatura de extração (2), volume de amostra (3) e concentração de eletrólito (4) como importantes para o sistema;logo as mesmas foram avaliadas em mais níveis pela matriz Doehlert.As variáveis 1, 2 e 3 foram apontadas como as que influenciam o sistema. O modelo quadrático do planejamento Doehlert foi significativo não apresentando falta de ajuste e a condição ótima foi definida no tempo e temperatura de extração de 60 min e 85 °C, respectivamente, com um volume de amostra de 11 mL e 0,04 mol/L de concentração de eletrólito. O método otimizado foi validado considerando os parâmetros seletividade, linearidade, precisão (repetitividade e precisão intermediária), exatidão e os limites de detecção (LOD) e quantificação (LOQ). O LOQ e LOD foram 0,76 e 2,28 μg/L, respectivamente. O método se mostrou linear com R2 igual a 0,992. Os resultados indicaram que o método HS-SPME/GC-ECD proposto é eficiente para a determinação de clorpirifós em água e as análises das amostras coletadas indicaram a presença de clorpirifós em concentrações entre osvalores de LOD e LOQ. Os resultados desta pesquisa foram informados à Prefeitura Municipal de Ouro Branco e órgãos competentes. __________________________________________________________________________________________ / ABSTRACT: Produced since1940, organophosphates were the first to replace the organochlorines, which already had bug resistance. Among these chlorpyrifos presents itself as a highly toxic insecticide. Due to its high lipossolubility it is absorbed by the human body, especially the skin. Organophosphates are known to induce or worsen certain health problems; so it is important its monitoring. For the determination of chlorpyrifos in aqueous samples generally are used steps of extraction/pre-concentration before performing the measurement. In this study it was investigated the liquid-liquid extraction technique withlow temperature partition (LLE-LTP) and solid phase microextraction with headspace (HS-SPME) and the techniques of gas chromatography with electron capture detection (GC- ECD) and gas chromatography coupled to mass spectrometry (GC-MS) to define the most sensitive analytical method, using multivariate experimental designs for the optimization of the sample preparation techniques. After optimization, the HS-SPME/GC-ECD method was validated for the monitoring of chlorpyrifosin surface water bodies close to the main potato fields in countryside of OuroBranco/MG; the presence of pesticides, such as chlorpyrifos, in these streams has been linked in previous research to the high rates of hypertension and abortion cases of the population in rural areas. The definition of the sampling was made after field visits and information survey obtained from the competent agencies and people. The full factorial design showed the variables time (1) and temperature of extraction (2), sample volume (3) and concentration of electrolyte (4) as important to the system; in this way the same ones were evaluated in more levels by Doehlert matrix. Variables 1, 2 and 3 were identified as those that affect the system, the quadratic modelof Doehlert design was significant not showing lack of fit and the optimal condition was set in the time and temperature of extraction of 60 min and 85 °C, respectively, with a volume of 11 mL and 0.04 mol/L of electrolyte concentration. The optimized method was validated considering the parameters selectivity, linearity, precision (repeatability and intermediate precision), accuracy and limits of detection (LOD) and quantitation (LOQ). The LOD and LOQ were 0.76 and 2.28 μg/L, respectively. The method was linear with R2 equal to 0.992. The results indicate that the method HS-SPME/ECD-GC proposed is efficient for the determination of chlorpyrifos in water and the analysis of the samples indicated the presence of chlorpyrifos in concentrations between the values of LOD and LOQ. These results were reported to the Ouro Branco City Hall and competent agencies.