No / The rate at which toxicological data is generated is continually becoming more rapid and the volume of data generated is growing dramatically. This is due in part to advances in software solutions and cheminformatics approaches which increase the availability of open data from chemical, biological and toxicological and high throughput screening resources. However, the amplified pace and capacity of data generation achieved by these novel techniques presents challenges for organising and analysing data output. Big Data in Predictive Toxicology discusses these challenges as well as the opportunities of new techniques encountered in data science. It addresses the nature of toxicological big data, their storage, analysis and interpretation. It also details how these data can be applied in toxicity prediction, modelling and risk assessment.

<p dir="ltr">Aquatic ecosystems and their inhabitants are no strangers to stressors. Natural stressors like disease, competition, and predation have a near constant presence in these environments and are often accompanied by human-induced stressors like climate change and chemical contaminants. Chemical contaminants like pesticides are often found in aquatic systems located near agriculture and can have detrimental effects on wildlife. Although natural stressors and pesticides often occur at the same time, their combined interactions still require further investigation to understand, as some pesticides, like fungicides, are frequently under researched. Additionally, fungicides also are lacking in research pertaining to combined chemical stressors. Studying the interactive effects of combined stressors, whether natural or human-induced, is crucial for applying laboratory findings to natural environments.</p><p dir="ltr">Here, I conducted an outdoor mesocosm experiment and multiple laboratory experiments to broadly assess the interactive effects of the fungicide chlorothalonil. More specifically, I explored 1) the interactive effects of the herbicide atrazine and chlorothalonil on an aquatic community, 2) the interaction between chlorothalonil and predator-induced stress of three tadpole species, and 3) the impacts of chlorothalonil on amphibian predator-prey dynamics using tadpoles and larval salamanders. First, to test my hypothesis that atrazine and chlorothalonil will have combined interactions that negatively impact an aquatic community, I conducted a mesocosm experiment where I exposed an aquatic community to atrazine, chlorothalonil, and the combination of the two pesticides. I found that the two pesticides do hold the potential to interact in certain cases, but their main effects alone are often just as damaging, especially for chlorothalonil where an environmentally relevant concentration caused near total morality for bullfrog tadpoles.</p><p dir="ltr">Next, I conducted a laboratory experiment with tadpoles and caged predators exposed to a sublethal concentration of chlorothalonil to test my hypothesis that the combination of predator-induced stress and chlorothalonil will decrease tadpole survival and alter tadpole behavior. I found that chlorothalonil alters tadpole behavior by significantly reducing activity levels in all three study species. However, no evidence was found for the interaction of the two stressors decreasing survival.</p><p dir="ltr">Lastly, to test my hypothesis that the toxicity of chlorothalonil will reduce tadpole behavioral responses and thereby increase their predation risk, I conducted a laboratory experiment with free-swimming tadpoles as prey and larval tiger salamanders as predators, exposed to different combinations of chlorothalonil exposure for the prey or predator. I found no meaningful differences in survival curves for the different combinations of prey and predator, but leopard frog tadpole final survival was lower in the higher chlorothalonil dose treatment due to predation.</p><p dir="ltr">Overall, these studies have assisted in filling research gaps on fungicides and their impact on predator-prey dynamics and aquatic communities. This work provided insights into the direct impacts of the herbicide atrazine and the fungicide chlorothalonil on aquatic species, and their potential to interact with natural stressors, emphasizing the need to protect natural ecosystems from chemical stressors.</p>

Single species toxicity tests are widely used to assess the potential effects of a toxicant on aquatic life. Increasingly, it is necessary to understand how the results of these tests relate to toxicant effects in natural communities. This dissertation presents the methodology and validation for a community level toxicity test that bridges the gap between single species tests and natural community responses. The research involved control of environmental parameters, improvement of feeding regimes and testing of the final community. The results are presented as four separate papers. The first paper addresses the development and validation of a standardized reconstituted water for culturing and toxicity testing of algae, cladocerans, a rotifer and two fish species. The next two papers address the substitution of the food source currently used in the fathead minnow survival-growth impairment test (<i>Artemia</i>) with a freshwater food source (the rotiter, <i>Brachionus calyciflorus</i>). Along with the alga <i>Chlorella vulgaris</i> (producer), B. calyciflorus (primary consumer) and the fathead minnow larvae (secondary consumer) comprise a three level food chain that was used to address trophic level interactions (feeding reduction and growth impairment) in the final phase of this research. The end result is an experimental procedure in which environmental parameters (water quality, temperature, etc.) and trophic structure parameters (Le. producer and primary consumer density) can be controlled well enough to insure that any shifts in community structure can be attributed to toxicant related effects. / Ph. D.

The purpose of the investigation presented in chapter 1 was to determine which of the following artificial stream designs would be most logistically simple yet effective in maintaining riffle insects during a 30-d bioassay: 1) static and no current (S-NC); 2) flow-through and no current (FT-NC); 3) static with current (S-C); or 4) flow-through with current (FT-C). Flow-through and current, when provided, were 12 ml min⁻¹ and 30 cm sec⁻¹, respectively. Streams were covered by emergence traps, and daylight equivalent lights provided a natural photoperiod. The four stream designs were evaluated in triplicate based on changes in insect species-abundances after 30 d. Test organisms were transferred to the artificial streams in rock-filled containers previously colonized for 30 d in a third-order mountain stream riffle. Relative to benthic samples taken directly from the source riffle, the artificial substrates selected for collector-filterers and against collector-gatherers. The FT-C and S-C stream designs maintained most taxa at or above initial densities. Emergent adults comprised a large proportion of mayfly and chironomid densities and must be monitored during bioassays with aquatic insects. The Investigation reported in chapter 2 was conducted to determine if contaminant-induced changes in macroinvertebrate and periphyton communities in laboratory stream microcosms could be used to predict macroinvertebrate and periphyton responses In a natural stream receiving the same contaminant. The microcosms were dosed in quadruplicate with four (0.0, 0.1, 1.0, and 10.0%) concentrations of a complex effluent; these concentrations reflected those in the field. Mayfly densities in the microcosms were significantly (P≤0.05) reduced at 1.0 or 10.0% effluent depending on species. Hydropsychlds were not affected by the effluent, and chironomids and periphyton were stimulated. Overall, the stream microcosms accurately predicted the macroinvertebrate and periphyton response observed in the field. Chapter 3 compared responses to a complex effluent from microcosms of indigenous macroinvertebrates and protozoans to responses observed in acute tests with Daphnia magna, Ceriodaphnia dubia and Pimephales promelas and chronic survival and reproductive tests with C. dubia The predictive utility of these various tests was then evaluated against observed effects in the receiving stream. The LC₅₀_s (% effluent) from the acute tests were 63.09 for Pimephales promelas, 18.8 to 31.3 for Daphnia magna and 54.7 for Ceriodaphnia dubia. Results from 7-day chronic tests indicated that C. dubia survival was significantly (P≤0.05) affected at 30% effluent and reproduction was affected at concentrations ≥3.0% effluent. In the protozoan microcosms, community composition was significantly (P≤0.05) changed at 1.0%; while protozoan species richness was significantly reduced at 3.0% effluent. The microcosms not only were the most sensitive indicators of effluent toxicity, they also correctly predicted which indigenous organisms would be lost and which would be stimulated at various ambient concentrations of the effluent. In the fourth chapter canonical discriminant analysis, 2 diversity indices, and 7 community comparison indices were evaluated to determine their utility in quantifying macroinvertebrate response to a complex effluent in laboratory microcosms. A permutation and randomization procedure was used to test the hypothesis of no treatment effect based on the community comparison indices. The Bray-Curtis index provided the most meaningful condensation of the data. / Ph. D. / incomplete_metadata

A avaliação ecotoxicológica dos sedimentos dos reservatórios de Barra Bonita e Salto Grande foi realizada através de testes de toxicidade aguda e crônica com o sedimento total e o solubilizado, com os organismos-teste Chironomus xanthus, Daphnia similis, Ceriodaphnia dubia, Vibrio fisheri e Spirillum volutans, além de análises físicas (pH, EH e \'O IND.2\') e químicas (fósforo, nitrogênio, carbono orgânico total, sulfetos e amônia e metais potencialmente disponíveis). A concentrações médias de matéria orgânica e fósforo obtidas para os sedimentos dos reservatórios de Barra Bonita e Salto Grande foram elevadas, e condizentes com a carga de matéria orgânica e com as baixas concentrações de oxigênio dissolvido e de potencial redox. Em relação aos metais, os resultados revelaram que, para o reservatório de Barra Bonita, eles podem estar influenciando a toxicidade detectada através dos testes de toxicidade. Em Salto Grande, os resultados indicaram que os sulfetos estão atuando como fase controladora dos metais, mantendo-os indisponíveis nos sedimentos. Para esse reservatório a principal causa da toxicidade foi a acidez. O estudo de Avaliação e Identificação da Toxicidade realizado com as amostras de sedimento dos reservatórios de Barra Bonita, Salto Grande e Rasgão mostrou que os possíveis compostos tóxicos responsáveis pela toxicidade são metais (Barra Bonita); amônia (Salto Grande e Rasgão); compostos ácidos voláteis e compostos orgânicos não iônicos (Rasgão). O tratamento com a resina zeolita foi eficiente na remoção da amônia e redução da toxicidade do sedimento do reservatório de Rasgão, confirmando dessa forma, a amônia como o principal composto tóxico. O estudo de Avaliação e Identificação da Toxicidade deve ser incorporado nos programas de controle, monitoramento e gestão dos recursos hídricos, devido a importância da identificação dos compostos responsáveis pela toxicidade. / An ecotoxicological evaluation of the sediments of Barra Bonita and Salto Grande reservoirs was performed by carrying out acute and chronic toxicity tests with both the total sediment and elutriate using Chironomus xanthus, Daphnia similis, Ceriodaphnia dubia, Vibrio fisheri and Spirillum volutans as test-organisms, as well as physical (pH, EH e \'O IND.2\') and chemical analysis (phosphorus, nitrogen, total organic carbon, sulphide, ammonium and potentially available metals). The mean concentrations of organic matter and phosphorus obtained for the sediments of Barra Bonita and Salto Grande reservoirs were high and in agreement with the organic matter loading, low dissolved oxygen concentrations and redox potential. In relation to metals, the results revealed that in Barra Bonita reservoir they might be responsible for the toxicity detected. For Salto Grande, the results indicated that sulphides act as metal controlling phase keeping them unavailable in the sediments. The main cause of the toxicity in this reservoir was acidity. The Toxicity Identification and Evaluation (TIE) carried out with the sediments of Barra Bonita, Salto Grande and Rasgão have shown that the probable toxic compounds responsible for toxicity are metals (Barra Bonita); ammonium (Salto Grande and Rasgão); volatile acid compounds and non-ionic compounds (Rasgão). The treatment with zeolite was efficient in removing ammonium and reducing toxicity in the sediment of Rasgão reservoir, thus confirming ammonium as the main toxic compound. The Toxicity Identification and Evaluation approach must be incorporated in the control, monitoring and management of water resources, due to the relevance of identifying the compounds responsible by the toxicity.

A indústria do petróleo possui extensa variedade de riscos ocupacionais em virtude da diversidade de atividades necessárias, seja na extração em águas profundas ou em outros locais isolados como em florestas e desertos, seja no processo de refino cada vez mais assemelhado à petroquímica. Para tanto, os trabalhadores desta indústria possui praticamente todos os riscos ocupacionais, ou seja, físico, químico, biológico. A base energética da sociedade atual é o petróleo não havendo fonte de energia alternativa economicamente viável para a sustentação da economia e indústria nos dias atuais, portanto, mecanismos eficazes de gerenciamento do risco devem ser perseguidos continuamente. Com avanço da higiene ocupacional, legislações modernas e restritivas aos riscos, a exposição ocupacional deste setor tem diminuído de forma bastante representativa nos últimos anos, e valores de limites de tolerância de várias substâncias cada vez menores tem obrigado à equipe de saúde ocupacional a desenvolverem estudos mais aprimorados. Conceitos de biomarcadores de exposição há muito é utilizado, porém falta definição de parâmetros para avaliação do Sistema Nervoso. Vários estudos indicam que exposições a baixas concentrações podem acarretar em dano à saúde, especialmente aos indivíduos que possuem uma susceptibilidade aumentada. Outro fator que merece atenção é a exposição a baixas concentrações de múltiplos agentes concomitantemente, que pode acarretar em efeito sinérgico para agressão aos mais variados órgãos e sistemas e para isso são necessários os biomarcadores de efeitos. Este trabalho visa discutir o papel dos biomarcadores de efeito precoce de neurotoxicidade de modo a preservar a saúde e qualidade de vida dos trabalhadores envolvidos no processo de produção. / The Oil industry has activities in forests, seas, deserts, cities, trough the extraction and transportation of crude oil and posterior transformation into products for consumers. This transformation occurs trough refinement Then people who work in oil companies are submitted to all of the occupational\'s risks, that is, physic , chemic and biological. In our society there is no economically practicable alternative source of energy yet, to support economy and industry, therefore efficient mechanisms of risk\'s management should be always pursuited. The improvement of legislation and occupational\'s hygiene have been reducing contact with chemist\'s products in last years and lesser Threshold Limit Values has been encouraging the occupational\'s health staff to develop better controls. Biomarker\'s conceptions are being used for a long time, but there is a lack of standards\' definition to evaluate the nervous system. Many papers indicate that exposures to low concentrations can cause health injury specially in individuals that have an increased susceptibility. Another fact that deserves attention is the exposure to low concentrations of multiple chemists\' substances concomitantly, which can cause an enlargement of the aggression of many organs and systems. The goal of this study is to discuss the role of the biomarkers of neurotoxicities, to preserve health and quality of life of the individuals who work at the production\'s process.

A boldina é o principal alcaloide encontrado no Peumus boldus, muito utilizado na medicina tradicional principalmente pelos seus efeitos gastrointestinais e hepáticos. Estudos indicam que apresenta diversas atividades farmacológicas como coletérica, hipnótica, citoprotetora, antitumoral, antiinflamatória, antipirética, antiplaquetária, antiplasmódica, antidiabética, antihipertensiva, inibidora da tirosinase e da acetilcolinesterase, antagonista dopaminérgico (receptores D1 e D2 like), adrenérgico (receptores α1 e α2) e serotoninérgico (receptores 5-HT3), entre outras. O chá de boldo (Peumus boldus) é muito utilizado por gestantes para aliviar os efeitos negativos da gestação como, constipação, cólicas uterinas, cefaleia e náuseas; controverso a esse uso ele também é muito utilizado como abortivo. Este estudo procurou identificar qual sua ação em ratas Wistar tratadas durante o período gestacional, GD1 ao dia anterior ao parto, nas doses de 1 mg/kg, 10 mg/kg e 100 mg/kg, uma vez ao dia, no ciclo claro, por meio gavagem. Conclui-se que o uso da boldina e do chá de P. boldus deve ser evitado durante o período gestacional pois seu uso na gestação causou alterações no desempenho reprodutiva das fêmeas, perda pré e pós implantação e mortes ao nascimento e pós-natais, além da modificação do comportamento materno (latência para lamber); foram encontradas também alterações no desenvolvimento da proles de fêmeas tratadas com boldina, em seu desempenho nos testes de reflexos (teste de endireitamento, geotaxia e agarrar), físico (abertura de olhos) e reprodutivo (abertura vaginal, descida de testículos e separação prepucial), assim como alteração no peso de órgãos de machos e fêmeas, na produção de espermatozoides e no percentual de espermatozoides com alteração (anormais). / Boldine is the main alkaloid found in Peumus boldus, widely used in traditional medicine mainly for its gastrointestinal and hepatic effects. Studies indicate that it presents several pharmacological activities as a choleretic, hypnotic, cytoprotective, anti-inflammatory, anti-plasmodic, antipyretic, antiplatelet, antidiabetic, antihypertensive, tyrosinase and acetylcholinesterase inhibitor, dopaminergic antagonist (D1 and D2 like receptors), α-adrenergic receptors antagonist (α 1 and α 2 receptors) and serotonergic receptors antagonist (5-HT 3 receptors), among others. Boldo’s tea (Peumus boldus) is widely used by pregnant women to alleviate the negative effects of pregnancy such as constipation, uterine cramps, headache and nausea; Controversely it is also widely used as an abortive substance The aim of this study was to identify the effect of boldine on Wistar rats treated during the gestational period, day 1 of gestation to the day before birth, at doses of 1mg/kg, 10mg/kg and 100mg/kg, daily, light cycle, by gavage. We concluded that the use of boldine and P. boldus tea should be avoided during the gestational period because its use in the gestacional period caused alterations in the reproductive performance of females, loss of pre and post implantation and deaths at birth and postnatal, besides modification of maternal behavior (latence to pup licking); alterations in the development of the offspring which the females were treated with boldine were observed, in their performance in the reflex tests (straightening, geotaxy and grab), physical development (eye opening) and reproductive development (vaginal opening, testicle descent and preputial separation), as well as changes in the weight of male and female organs, in the production of spermatozoa and in the percentage of abnormal spermatozoa.

Wong Chun-kwan. / Thesis submitted in: December 1999. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 127-137). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgments --- p.viii / Tables of Contents --- p.ix / List of Figures --- p.xv / List of Tables --- p.xxvi / Chapter Chapter 1: --- INTRODUCTION --- p.1 / Chapter Chapter 2: --- LITERATURE REVIEW --- p.8 / Chapter 2.1 --- Toxicology --- p.8 / Chapter 2.1.1 --- Acute toxicity test --- p.8 / Chapter 2.1.2 --- Biochemical Analysis --- p.9 / Chapter 2.1.3 --- Organ weights --- p.10 / Chapter 2.2 --- Histology --- p.11 / Chapter 2.2.1 --- Light Microscope --- p.11 / Chapter 2.2.2 --- Electron Microscopy --- p.11 / Chapter 2.3 --- Tissue injury --- p.12 / Chapter 2.3.1 --- Free-radical mechanisms --- p.12 / Chapter 2.3.2 --- Lipid peroxidation --- p.13 / Chapter 2.4 --- Carbon tetrachloride (CC14) --- p.14 / Chapter 2.4.1 --- Mechanisms of carbon tetrachloride toxicity --- p.15 / Chapter 2.5 --- Trichloroethylene (TCE) --- p.18 / Chapter 2.5.1 --- Mechanisms of trichloroethylene toxicity --- p.21 / Chapter 2.6 --- Dimethyl sulfoxide (DMSO) --- p.25 / Chapter 2.7 --- N-acetylcysteine (NAC) --- p.27 / Chapter Chapter 3: --- MATERIALS AND METHODS --- p.28 / Chapter 3.1 --- Materials --- p.28 / Chapter 3.2 --- Methods --- p.31 / Chapter 3.2.1 --- Acute hepatotoxicity test on aqueous seaweed extracts --- p.31 / Chapter 3.2.1.1 --- Preparation of aqueous extracts of seaweed --- p.31 / Chapter 3.2.1.2 --- Experimental protocol --- p.31 / Chapter 3.2.1.3 --- Biochemical assays --- p.32 / Chapter 3.2.1.4 --- Organ weights --- p.36 / Chapter 3.2.1.5 --- Histopathological examination --- p.36 / Chapter 3.2.1.6 --- Statistical analysis --- p.36 / Chapter 3.2.2 --- Curative and preventive tests of seaweed aqueous extracts against the CCl4-induced hepatotoxicity --- p.37 / Chapter 3.2.2.1 --- Preparation of aqueous extracts of seaweed --- p.37 / Chapter 3.2.2.2 --- Experimental protocol --- p.37 / Chapter 3.2.2.3 --- Biochemical assays --- p.39 / Chapter 3.2.2.4 --- Organ weights --- p.39 / Chapter 3.2.2.5 --- Histopathological examination --- p.40 / Chapter 3.2.2.6 --- Statistical analysis --- p.41 / Chapter 3.2.3 --- Acute hepatotoxicity test of TCE in rats by oral and intraperitoneal routes --- p.42 / Chapter 3.2.3.1 --- Experimental protocol --- p.42 / Chapter 3.2.3.2 --- Biochemical assays --- p.43 / Chapter 3.2.3.3 --- Organ weights --- p.43 / Chapter 3.2.3.4 --- Histopathological examination --- p.44 / Chapter 3.2.3.5 --- Statistical analysis --- p.44 / Chapter 3.2.4 --- Curative and preventive tests of seaweed aqueous extracts against the TCE effective dose-induced toxicity --- p.44 / Chapter 3.2.4.1 --- Preparation of aqueous extracts of seaweed --- p.44 / Chapter 3.2.4.2 --- Experimental protocol --- p.45 / Chapter 3.2.4.3 --- Biochemical assays --- p.46 / Chapter 3.2.4.4 --- Organ weights --- p.46 / Chapter 3.2.4.5 --- Histopathological examination --- p.46 / Chapter 3.2.5 --- Antidotal effects of dimethyl sulfoxide (DMSO) and N-acetylcysteine (NAC) against CC14- and TCE- induced poisoning in rats --- p.47 / Chapter 3.2.5.1 --- Experimental protocol --- p.47 / Chapter 3.2.5.2 --- Biochemical assays --- p.48 / Chapter 3.2.5.3 --- Organ weights --- p.48 / Chapter 3.2.5.4 --- Histopathological examination --- p.49 / Chapter 3.2.6 --- Hepatoprotective effect of seaweeds' methanol extract against CC14- and TCE-induced poisoning in rats --- p.49 / Chapter 3.2.6.1 --- Preparation of methanol extracts of seaweed --- p.49 / Chapter 3.2.6.2 --- Experimental protocol --- p.50 / Chapter 3.2.6.3 --- Biochemical assays --- p.52 / Chapter 3.2.6.4 --- Organ weights --- p.52 / Chapter 3.2.6.5 --- Histopathological examination --- p.53 / Chapter Chapter 4 --- RESULTS --- p.54 / Chapter 4.1 --- Acute hepatotoxicity test on aqueous seaweed extracts --- p.54 / Chapter 4.1.1 --- The biochemical assays of the serum transaminase activity --- p.54 / Chapter 4.1.2 --- The organ weight (Aqueous seaweed crude extracts) --- p.56 / Chapter 4.2 --- Curative and preventive tests of seaweed aqueous extracts against the CCl4-induced hepatotoxicity --- p.58 / Chapter 4.2.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.58 / Chapter 4.2.2 --- The organ weight (Curative) --- p.60 / Chapter 4.2.3 --- The biochemical assays of the serum transaminase activity (Preventive) --- p.62 / Chapter 4.2.4 --- The organ weight (Preventive) --- p.64 / Chapter 4.3 --- Acute hepatotoxicity test of TCE in rats by oral and intraperitoneal routes --- p.66 / Chapter 4.3.1 --- Oral route --- p.66 / Chapter 4.3.1.1 --- One-time oral route --- p.66 / Chapter 4.3.1.2 --- Two-time oral route --- p.66 / Chapter 4.3.2 --- Intraperitoneal route --- p.66 / Chapter 4.3.3 --- Time course of the effective dose of 20% TCE in i.p. route --- p.67 / Chapter 4.4 --- Curative and preventive tests of seaweed aqueous extracts against the TCE effective dose-induced toxicity --- p.12 / Chapter 4.4.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.72 / Chapter 4.4.2 --- The organ weight (Curative) --- p.74 / Chapter 4.4.3 --- The biochemical assays of the serum transaminase activity (Preventive) --- p.76 / Chapter 4.4.4 --- The organ weight (Preventive) --- p.78 / Chapter 4.5 --- Antidotal effects of dimethyl sulfoxide (DMSO) and N-acetylcysteine (NAC) against CC14- and TCE-induced poisoning in rats --- p.80 / Chapter 4.5.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.80 / Chapter 4.5.2 --- The organ weight (Curative) --- p.82 / Chapter 4.5.3 --- The biochemical assays of the serum transaminase activity (Preventive) --- p.84 / Chapter 4.5.4 --- The organ weight (Preventive) --- p.86 / Chapter 4.6 --- Hepatoprotective effect of methanol extract of seaweed against CC14- and TCE-induced poisoning in rats --- p.88 / Chapter 4.6.1 --- The biochemical assays of the serum transaminase activity (Curative) --- p.88 / Chapter 4.6.2 --- The organ weight (Curative) --- p.89 / Chapter 4.7 --- Histopathological examinations --- p.90 / Chapter 4.7.1 --- Acute hepatotoxicity test on aqueous seaweed extracts --- p.91 / Chapter 4.7.2 --- Curative and preventive tests of seaweed aqueous extracts against the CC14-induced hepatotoxicity --- p.92 / Chapter 4.7.3 --- Acute hepatotoxicity test of TCE in rats by oral and intraperitoneal routes --- p.99 / Chapter 4.7.4 --- Curative and preventive tests of seaweed aqueous extracts against the TCE effective dose-induced toxicity --- p.100 / Chapter 4.7.5 --- Antidotal effects of dimethyl sulfoxide (DMSO) and N-acetylcysteine (NAC) against CC14- and TCE-induced poisoning in rats --- p.100 / Chapter 4.7.6 --- Hepatoprotective effect of methanol extract of seaweed against CC14- and TCE-induced poisoning in rats --- p.102 / Chapter Chapter 5 --- DISCUSSION --- p.106 / Chapter Chapter 6 --- CONCLUSION --- p.124 / REFERENCES --- p.127 / APPENDIX --- p.138

Leung Bo-Shan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (leaves 148-159). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.v / List of Abbreviations --- p.vi / List of Tables --- p.viii / List of Figures --- p.ix / Chapter 1 --- Introduction / Chapter 1.1 --- Food colorants --- p.1 / Chapter 1.2 --- Caramel --- p.3 / Chapter 1.2.1 --- Classes of caramel --- p.3 / Chapter 1.2.2 --- Toxicological studies of caramel --- p.5 / Chapter 1.3 --- Castanea mollissima --- p.9 / Chapter 1.4 --- Antioxidants --- p.10 / Chapter 1.4.1 --- Background --- p.10 / Chapter 1.4.2 --- Methods used to evaluate the antioxidative activity --- p.12 / Chapter 1.4.2.1 --- DPPH* scavenging method --- p.13 / Chapter 1.4.2.2 --- High performance liquid chromatography (HPLC) --- p.13 / Chapter 1.5 --- Microtox® test --- p.19 / Chapter 1.6 --- Mutatox® test --- p.19 / Chapter 1.7 --- Methods used to evaluate the functions of major organs --- p.20 / Chapter 1.7.1 --- Liver --- p.20 / Chapter 1.7.2 --- Kidneys --- p.23 / Chapter 1.8 --- Toxicology --- p.25 / Chapter 1.8.1 --- Acute toxicity test --- p.25 / Chapter 1.8.2 --- Chronic toxicity test --- p.26 / Chapter 1.9 --- Objective --- p.27 / Chapter 2 --- Materials and Methods --- p.28 / Chapter 2.1 --- Plant materials --- p.28 / Chapter 2.2 --- Sample preparation --- p.28 / Chapter 2.3 --- Pigment characterization --- p.30 / Chapter 2.3.1 --- Stability test --- p.30 / Chapter 2.3.2 --- HPLC separation of CP --- p.31 / Chapter 2.3.3 --- Determination of antioxidative activity with the DPPH* scavenging method --- p.31 / Chapter 2.4 --- Microtox® test --- p.33 / Chapter 2.5 --- Mutatox® test --- p.34 / Chapter 2.6 --- Acute toxicity test --- p.35 / Chapter 2.6.1 --- Animals --- p.35 / Chapter 2.6.2 --- Housing and maintenance --- p.35 / Chapter 2.6.3 --- Experimental design --- p.37 / Chapter 2.6.4 --- Chemicals --- p.39 / Chapter 2.6.5 --- Clinical pathology test --- p.41 / Chapter 2.6.5.1 --- Haematology --- p.41 / Chapter 2.6.5.2 --- Blood chemistry --- p.45 / Chapter 2.6.5.3 --- Urinalysis --- p.55 / Chapter 2.6.6 --- Histological study --- p.57 / Chapter 2.6.7 --- Statistical analysis --- p.57 / Chapter 2.7 --- Chronic toxicity test --- p.59 / Chapter 2.7.1 --- Animals --- p.59 / Chapter 2.7.2 --- Housing and maintenance --- p.59 / Chapter 2.7.3 --- Experimental design --- p.59 / Chapter 2.7.4 --- Chemicals --- p.60 / Chapter 2.7.5 --- Clinical pathology test --- p.61 / Chapter 2.7.5.1 --- Haematology --- p.61 / Chapter 2.7.5.2 --- Blood chemistry --- p.62 / Chapter 2.7.5.3 --- Urinalysis --- p.62 / Chapter 2.7.6 --- Histological study --- p.62 / Chapter 2.7.7 --- Statistical analysis --- p.62 / Chapter 3 --- Results --- p.63 / Chapter 3.1 --- Pigment characterization --- p.63 / Chapter 3.1.1 --- Stability test --- p.63 / Chapter 3.1.2 --- HPLC separation of CP --- p.63 / Chapter 3.1.3 --- Antioxidative activities of CP preparations --- p.63 / Chapter 3.2 --- Microtox® test --- p.65 / Chapter 3.3 --- Mutatox® test --- p.65 / Chapter 3.4 --- Acute toxicity test --- p.66 / Chapter 3.4.1 --- Growth rate --- p.66 / Chapter 3.4.2 --- Food and fluid consumption --- p.66 / Chapter 3.4.3 --- Organ-weight --- p.66 / Chapter 3.4.4 --- Clinical pathology tests --- p.68 / Chapter 3.4.4.1 --- Haematology --- p.68 / Chapter 3.4.4.2 --- Blood chemistry --- p.70 / Chapter 3.4.4.3 --- Urinalysis --- p.76 / Chapter 3.4.5 --- Histological study --- p.76 / Chapter 3.5 --- Chronic toxicity test --- p.77 / Chapter 3.5.1 --- Growth rate --- p.77 / Chapter 3.5.2 --- Food and fluid consumption --- p.77 / Chapter 3.5.3 --- Organ-weight --- p.77 / Chapter 3.5.4 --- Clinical pathology tests --- p.78 / Chapter 3.5.4.1 --- Haematology --- p.78 / Chapter 3.5.4.2 --- Blood chemistry --- p.80 / Chapter 3.5.4.3 --- Urinalysis --- p.82 / Chapter 3.5.5 --- Histological study --- p.82 / Chapter 4 --- Discussion --- p.137 / Chapter 4.1 --- Pigment characterization --- p.137 / Chapter 4.2 --- Toxicological studies of CP --- p.140 / Chapter 5 --- Conclusion --- p.147 / References --- p.148

O gênero Drimys é o de maior área de distribuição geográfica da família (Winteraceae), que compreende sete gêneros e cerca de 120 espécies. No Brasil, encontra-se desde a Bahia até o Rio Grande do Sul e ocorre em duas espécies, Drimys angustifolia Miers. e Drimys brasiliensis Miers, conhecidas popularmente como “casca-de-anta”. As folhas e cascas são usadas na medicina popular como antiescorbútico, estimulante, antiespasmódica, antidiarreica, antifebril, contra hemorragia uterina, antibacteriana, no tratamento de asma, bronquite, certas afecções do trato digestivo e, algumas vezes no tratamento do câncer e inseticidas. Ambas são caracterizadas pela presença de flavonóides e sesquiterpenoides. O objetivo deste trabalho foi o de avaliar a composição química, o potencial antioxidante, antiviral, inseticida, antitumoral, antifúngico, avaliação tóxica aguda e, também, formular e caracterizar duas nanoemulsões dos óleos voláteis das duas espécies do estudo. Por meio da hidrodestilação, foi possível realizar a extração dos óleos essenciais e determinar seus rendimentos que variaram entre 0,3 e 0,5% de folhas. A identificação dos componentes foi realizada utilizando a Cromatrografia Gasosa acoplada ao espectrofotômetro de massas (CG-EM), as quais apresentaram como componentes majoritários em todas as análises, o biciclogermacreno para a espécie Drimys angustifolia e a ciclocolorenona para a espécie Drimys brasiliensis. A formulação e caracterização da nanoemulsão foi realizada dentro dos parâmetros estabelecidos (tamanho de partícula, índice de polidispersão, pH) e através da microscopia eletrônica de transmissão foi confirmado o tamanho na escala nano. No que tange a atividade inseticida, ambas as espécies mostraram-se promissoras repelindo o cupim da espécie Cryptotermes brevis com índices de repelência negativos. Para avaliação inicial do potencial larvicida de Aedes aegypti, os óleos das duas espécies não foram eficientes. No entanto, a nanoemulsão de Drimys brasiliensis levou a 69% e 89% de mortalidade, correspondendo às concentrações de 0,5 μg/mL e 0,83 μg/mL, respectivamente. Com relação a atividade antioxidante, os óleos das duas espécies não foram capazes de reduzir o radical (2,2-difenil-1-picril-hidrazil) DPPH . Já na atividade antiviral frente ao VHS-1, as duas espécies mostraram-se promissoras inibindo a replicação viral. Na atividade antitumoral nas linhagens de glioma (U-138 MG) e de bexiga (T24), somente o óleo de Drimys brasiliensis reduziu a viabilidade celular das duas linhagens pelo ensaio do MTT, com resultados similares na contagem celular. Na análise da citometria pela incorporação da Anexina-V/Iodeto de propídio caracterizou-se indução de apoptose tardia. Na atividade antifúngica, as duas espécies foram eficazes contra o fungo oportunista Acremonium, presente em diversas infecções em transplantados e imunocomprometidos. Quanto à avaliação da toxicidade aguda, as duas espécies mostraram importantes sinais de toxicidade em ratos Wistar como ptose, tremor, redução da atividade motora, exolftamia, aumento da freqüência respiratória. Os animais tratados com espécie D. brasiliensis ainda apresentaram aumento na micção e diarréia. Desta maneira, é importante ressaltar que todos os ensaios biológicos, o estudo toxicológico, e a elaboração e caracterização das nanoemulsões dos óleos essenciais foram realizados pela primeira vez para as duas espécies vegetais, destacando a importância e ineditismo do presente trabalho. / The genus Drimys presents the largest geographical distribution of the family (Winteraceae), which comprises seven genera and about 120 species. In Brazil, is found from Bahia to Rio Grande do Sul and occurs in two species, Drimys angustifolia L. and Drimys brasiliensis Miers, popularly known as "Casca-de-anta". The leaves and barks are used in folk medicine as antiscorbutic, stimulant, antispasmodic, anti-diarrheal, antipyretic, against uterine bleeding, antibacterial, to treat asthma, bronchitis, certain disorders of the digestive tract and sometimes in the treatment of cancer and insecticides. Both are characterized by the presence of flavonoids and sesquiterpenoids. The objective of this study was to evaluate the chemical composition, the antioxidant, antiviral, insecticidal, antitumor, antifungal, acute toxicological evaluation and also to formulate and characterize two nanoemulsions of the volatile oils of two species of the study. By steam distillation was possible to perform the extraction of essential oils and to determine their yields ranging between 0.3 and 0.5% of leaves. The identification of components was performed using the Gas Cromatography coupled to mass spectrometer (GC-MS), which presented as major components in all analyzis bicyclogermacrene for the Drimys angustifolia species and ciclocolorenona for the Drimys brasiliensis. The formulation and characterization of nanoemulsion was performed within the established parameters (particle size, polydispersity, pH) and by transmission electron microscopy confirming the size at the nanoscale. Regarding the insecticidal activity, both species were promising repelling termites Cryptotermes brevis with negative index of repellency. To evaluate initial potential larvicidal Aedes aegypti, the oils of the two species were not efficient, however, of the Drimys brasiliensis nanoemulsion the larvae mortality of the 69% and 89%, corresponding to concentrations of 0.5 μg/mL and 0.83 μg/mL, respectively. With respect to antioxidant activity, the oils of the two species have not been able to reduce the (2,2-diphenyl-1-picrylhydrazyl) DPPH radical. In the antiviral activity against the HSV-1, the two species were promising inhibiting viral replication. In antitumor activity in glioma cell lines (U-138 MG) and bladder (T24), only the oil Drimys brasiliensis reduced cell viability of two strains by the MTT assay, with similar results in cell counts and flow cytometry analysis of the incorporation of Anexinn-V/Propidium Iodide characterized as late apoptosis. In the antifungal activity, the two species were effective against the opportunistic fungus Acremonium, present in various infections in transplant and immunocompromised. Regarding the evaluation of acute toxicity, the two species showed significant signs of toxicity in rats as ptosis, tremor, decreased motor activity, exolftamia, shortness of breath. Animals treated with species Drimys brasiliensis also showed an increase in urination and diarrhea. Thus, it is important that all biological assays, the toxicity study, and preparation and characterization of nanoemulsions of essential oils were first performed for the two species, highlighting the importance and originality of this work.