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Made available in DSpace on 2014-06-11T19:29:06Z (GMT). No. of bitstreams: 0 Previous issue date: 2003Bitstream added on 2014-06-13T20:38:17Z : No. of bitstreams: 1 aquino_ft_me_araiq.pdf: 385899 bytes, checksum: 3106d26f4f13b220d6c1372af6c103c2 (MD5) / Os ácidos orgânicos de cadeia curta: acético, propiônico, butírico e valérico tem sido descritos na literatura como sendo fitotóxicos. Esse estudo propõe o monitoramento desses ácidos durante o processo de compostagem de resíduos sólidos domésticos. Foram monitorados três tipos de leiras: leira com revolvimento (LR), leira com material de estrutura (LMe) e leira com cobertura (LC), todas com material de partida proveniente da Usina de Compostagem da cidade de Araraquara, São Paulo. Foram utilizadas três técnicas principais para o monitoramento desses ácidos: Cromatografia Gasosa (CG), Calorimetria Exploratória Diferencial (DSC) e testes de germinação em sementes. A cromatografia demonstrou que esses ácidos são encontrados em baixas concentrações e sua evolução foi curta, já que depois das primeiras amostragens, somente o ácido acético permaneceu, sendo que os demais não foram detectados ou estão abaixo de 5 μg.ml-1. O DSC mostrou-se interessante para avaliação do comportamento da evolução desses ácidos, já que foi apresentada uma influência da matriz sobre um dos analitos. Além disso, o DSC foi uma técnica complementar, que estabeleceu melhor estudo do comportamento do ácido propiônico. Os testes de germinação concluíram que esses ácidos não são fitotóxicos para concentrações de 5-500 μg.ml-1. Quanto à avaliação do composto, esse demonstrou-se apto para aplicação ao solo somente ao final do processo de compostagem. / The short chain organic acids: acetic, propionic, butyric and valeric has been reported in literature as been phytotoxic. This study propose the monitoring these acids during the composting process. It was monitored three types of solid waste dumps: only with turn over (LR); with structure material (LMe) and with a cover layer (LC), all of them with starter material from Composting Plant of Araraquara city, São Paulo. It was utilized three main techniques for monitoring these acids: Gas Chromatography (CG), Differential Scanning Calorimetry (DSC) and seed germination tests. The chromatography demonstrates these acids are present at low concentrations and your evolution was short, after first sample period only acetic acid remains, and the others weren’t detected, or are under 5 μg.ml-1. The DSC showed interesting for the evaluation of the occurence of these acids, that was showed a matrix influence on these analites. Besides, DSC was a complementar technique, that established a better study of propionic acid behaviour. The germination tests concluded these acids aren’t phytotoxic for the range: 5-500 μg.ml-1. To the compost evaluation, it has demonstrates that it was able to soil application only after end of the process.

Cis-diol-containing metabolites have attracted increasing attention in recent years. These metabolites widely exist in the body fluids and tissues. They play important roles in the structure, function and metabolic activity of cells. Some of them are related to cell proliferation and metabolic processes. And they have been used to denote a state of disease as potential biomarkers. Several methods have been developed for the analysis of cis-diol-containing metabolites. However, these methods faced a challenge to separate and detect isomers of these compounds, particularly for compounds with low abundance and high polarity. Therefore, novel methods were necessary to improve the separation and detection sensitivity of this kind of metabolites. With this aim, chemical derivatization methods were developed for cis-diol-containing metabolite detection by using liquid chromatography-mass spectrometry in this project. These methods were optimized and validated to achieve the optimal reaction conditions. And they were applied to study real-world biological systems, including the changes of modified nucleosides in hepatocellular carcinoma (HCC) nude mice and toxic effects of bisphenol A (BPA) exposure. Firstly, the derivatization reaction of cis-diol compounds with acetone were optimized. Factors that affected reaction efficiency were investigated by reacting guanosine (G) with acetone. The optimal reaction conditions were validated by detecting four acetonides of urinary nucleosides by using LC-MS/MS. The results showed that the approach had good linearity, accuracy and precision. The recoveries were ranged from 92.9% to 103.5%. It indicated that the assay was reproducible. The robust method should be potentially useful for the analysis of modified nucleosides and other cis-diol-containing metabolites in biological samples. The validated derivatization method was applied to determine urinary nucleosides by LC-MS. This method not only improved the retention of nucleosides on reversed-phase column, but also reduced the matrix effect from urine samples and enhanced detection sensitivity of mass spectrometry. Isotope labeling method with acetone-d6 and multivariate statistical analysis enabled the positive identification of 56 nucleosides, including 52 modified nucleosides. The obtained results indicated that the derivatization method was practical, fast and effective for the identification of urinary nucleosides. It was successfully applied to study the changes of urinary nucleosides in nude mice bearing HCC. Some significantly changed nucleosides were identified as potential biomarkers. Subsequently, this approach was modified by employing parallel reaction monitoring (PRM) method which was based on high resolution MS to detect urinary nucleosides in rats exposed to BPA. Comparing to the data acquired by triple quadrupole MS with neutral loss scanning, higher specificity and sensitivity were achieved by using PRM scanning mode. Therefore, more nucleosides were identified by using the method in urine samples (from 56 up to 66). The changes of the detected nucleosides were studied in the rats exposed to BPA. Various trends of modified nucleosides were observed with different dose BPA exposure. Specifically, the high-dose exposure group was the most strongly affected. The biomarker of RNA oxidation, 8-hydroxyguanosine (8-oxoG), showed significant change in this group. It proved that BPA exposure could induce RNA damage when the dose of BPA was beyond a certain amount. Except for nucleosides, other cis-diol-containing metabolites, such as carbohydrates, were also studied by using the derivatization method. Acetone and acetone-d6 were applied to label the cis-diol metabolites. Based on the chemical isotope labeling, cis-diol metabolites were easily recognized from urine samples. Influence of BPA exposure on these metabolites was investigated by comparing different doses of BPA administration on rats. Analytes showed noticeable difference were highlighted. Pathway analysis indicated that galactose metabolism, nucleoside and its analogues metabolism were disturbed. The derivatization method was extended to quantify nucleotides in plasma samples. According to the specific physical-chemical properties of nucleotides, the method was improved to fit the requirement of analysis by using 1,1-Dimethoxycyclohexane (DMCH) as derivatization agent and formic acid (FA) as catalyst. Tip micro-columns packed with TiO2 were used for selective adsorption of nucleotides in the plasma. Then in-situ derivatization were carried out to change the polarity of targeted compounds. LC-MS analysis of the derivatization products were employed without using ion-pairing reagents. This method exhibited a high selectivity for the extraction of nucleotides. After derivatization, retention of nucleotides on reversed-phase C18 column was improved. Complete separation of nucleotides with the same base was achieved. The peak shape was symmetrical and the tailing was eliminated by using high pH mobile phase. The method settled the problems of nucleotide detection, which were poor retention, trailing, in-source fragmentation and contamination of ion-pairing reagents. The quantitative method was successfully applied to determine the content of nucleotides in plasma samples of rats exposed to BPA. It was simple and fast, as well as good selectivity and stability. It could be extended to detection of other phosphorylated metabolites with similar structure. To our best knowledge, it was the first time to employ derivatization methods to detect cis-diol-containing metabolites. The methods decreased the matrix effects of complex biological samples, and also decreased the polarity of cis-diol-containing metabolites. The changes of properties not only improved the chromatographic separation, but also enhanced the MS intensities. The methods overcame the problems of cis-diol-containing metabolite detection on reversed-phase column. They were successfully applied to study the changes of cis-diol-containing metabolites of HCC and toxic effects of BPA exposure. The method might be extended to determine other cis-diol-containing metabolites in urine samples as well as in cells, tissues and plasma samples. It might be valuable for the understanding of the roles of cis-diol-containing metabolites in in cell metabolism.

Orientador: Caio Antonio Carbonari / Coorientador: Edivaldo Domingues Velini / Banca: Carlos Gilberto Raetano / Banca: Fernando Tadeu de Carvalho / Resumo: O sistema de produção com ou sem a pres ença de palhada, bem como, a aplicação de vinhaça nos canaviais, podem levar a alterações nas propriedades do solo e influenciar a disponibilidade dos herbicidas na solução do solo. Diante disto, o presente trabalho teve por objetivo avaliar a influência d a água, água após passar pela palha de cana - de - açúcar e vinhaça na disponibilidade na solução do solo dos herbicidas amicarbazone, clomazone, diuron, hexazinone, imazapic, sulfentrazone e tebuthiuron em diferentes solos. Adotou - se o esquema fatorial 3 x 31, com delineamento em blocos inteiramente casualizados com três repetições, sendo utilizadas 31 amostras de solos distintos quanto às características físicas e químicas e três tratamentos (água, água de lavagem de palha e vinhaça). As amostras de solo foram dispostas em bandejas e submetidas à aplicação dos herbicidas, em seguida foram acondicionadas em cartuchos plásticos e saturadas com água deionizada, água de lavagem de palha e vinhaça, permanecendo em repouso durante 18 horas sob refrigeração (8 ± 3 ° C) . Posteriormente, fez - se duas extrações, na primeira quantificou - se os herbicidas presentes na solução do solo e na segunda foi realizada a extração total do herbicida remanescente no solo para determinação da porcentagem de recuperação de cada herbicida t estado. Para as quantificações utilizou - se um sistema LC - MS/MS, composto por um Cromatógrafo Líquido de Alta Performance (HPLC) acoplado ao espectrômetro de massas. O amicarbazone e o imazapic foram mais 2 disponibilizados na solução dos solos com a aplicaçã o de água e água de lavagem de palha na maioria das amostras. Todavia, a adição de vinhaça promoveu uma maior disponibilização do diuron e tebuthiuron em todos os solos. O clomazone, hexazinone e sulfentrazone não apresentaram diferença significativa entre os ... / Abstract: The pr oduction system with or without the straw as how the sugarcane vinasse application may cause some changes in the soil properties and may influence the avaiability of the herbicides in the soil solution. Based on that, the objective of this study was to eva luate the water influence after passing throught the sugarcane straw and the vinasse, both correlated with the availability of the herbicides a micarbazone, clomazone, diuron, hexazinone, imazapc, sufentrazone and tebuthiuron in the soil solution in differ ent kinds of soils. The factorial 3 x 31 with complete randomized block design, with three replications, was used with 31 physical and chemical different soil samples and three treatments (water, wash water straw and vinasse). The soil samples were arrange d on trays and submitted to the herbicides application, then they were placed in plastic cartridges and satured with deionized water, wash water straw and vinasse, and remained resting for 18 hours under refrigeration (8 ± 3 o C) . After that, two extrations were made: the first one quantifying the presence of the herbicides in the soil solution and in the second one the total extraction of herbicide remaining in the soil was taken to determine the recovery percentage of each herbicide tested. For quantificat ions it was used a LC - MS/MS system, compound of a High Performance Liquid Chromatograph (HPLC) coupled with mass spectrometer. The amicarbazone and imazapic herbicides, in most samples, were more available in the soil solution with the water application an d in the wash water straw. However, vinasse had promoted an increase availability of diuron and tebuthiuron in all soils. Clomazone, hexazinone and sulfentrazone showed no significative difference between the treatments. The herbicides with the highest fre quency of availability of herbicides in soils were imazapic, hexazinone, amicarbazone and tebuthiuron. However, about 6% of ... / Mestre

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