Hidrocarbonetos aromáticos conhecidos como BTEX (benzeno, tolueno, etilbenzeno e xilenos) estão entre os maiores problemas de poluição ambiental. Estes compostos são reconhecidos por sua recalcitrância em ambientes anóxicos, e a remoção destes hidrocarbonetos, de sítios anaeróbios contaminados é dependente da atividade de uma população de microrganismos adaptados capazes de promover a biodegradação destes compostos, nestas condições. Neste sentido, o presente trabalho buscou purificar, caracterizar e utilizar cultura desnitrificante, como inóculo para desenvolvimento de biofilme, em reator anaeróbio de leito fixo preenchido com espumas de poliuretano, além de avaliar o potencial deste biofilme em promover a biodegradação dos compostos de BTEX. Células da cultura apresentaram coloração Gram negativa, com morfologia de cocos (diplococos e cocobacilos). A cultura foi capaz de crescer, sob condições desnitrificantes, utilizando diferentes substratos. A cultura não apresentou capacidade de crescer sob condições sulfetogênicas. Velocidades de crescimento ('mü') de 0,046/h e 0,050/h e tempos de geração (Tg) de 15,1 horas e 13,9 horas foram obtidos para as células crescidas em benzeno e etilbenzeno, respectivamente. A cultura purificada foi utilizada para formar biofilme em RAHLF. Em todas as condições de alimentação as quais os reatores foram submetidos houve remoção da matéria orgânica, nitrato e dos hidrocarbonetos. A menor eficiência de remoção de hidrocarbonetos foi de 89,4%, obtida durante a alimentação dos reatores com benzeno. A matéria orgânica (DQO) foi removida com eficiência média de 87,1%. A eficiência média de remoção de nitrato foi de 94%. O sequenciamento das amostras revelou que o biofilme do RAHLF1 foi formado, principalmente, por espécies de Paracoccus, Pseudomonas e Bacteroides, enquanto que no RAHLF2, alimentado com etilbenzeno, observou-se espécies dos gêneros: Paracoccus, Pseudomonas, Xanthomonas e Variovorax / The monoaromatic hydrocarbons known as BTEX (benzene, toluene, ethylbenzene, and xylene) are among the greatest environmental pollution problems. These compounds are known by their recalcitrance in anoxic environments, and the removal of these toxics from anaerobic contaminated sites depends on the presence of an adapted microbial population capable of to promote the biodegradation of these compounds under such condition. Thus, in the current study a denitrifying culture was purified, characterized and used as inoculum to form a biofilm in a horizontal-flow anaerobic immobilized biomass reactor filled with polyurethane foam, and to assess the potential of this biofilm to promote BTEX compounds biodegradation. Culture cells were Gram negative, with coccus (diplococcus and cocobacilli) morphology. The culture was able to grow, under denitrifying conditions, using different substrates. No growth was seen in sulfate-reducing conditions. Growth velocity('mü') of 0.046/h and 0.050/h, and generation time of 15.1 and 13.9 hours were obtained for cultures isolated in the presence of benzene and ethylbenzene, respectively. The purified culture was used to form a biofilm inside RAHLF. At all feeding conditions the reactors were submitted the organic matter, nitrate and hydrocarbons were removed. The smallest hydrocarbons removal efficiency was 89.4%, obtained during benzene feeding. The organic matter (COD) was removed with mean efficiency of 87.1%. The mean efficiency of nitrate removal was 94.0%. Sequencing of samples showed that the RAHLF1 biofilm was composed specially for Paracoccus, Pseudomonas and Bacteroides species. Biofilm of RAHLF2 reactor, obtained from ethylbenzene feeding, was composed by Paracoccus, Pseudomonas, Xanthomonas and Variovorax species

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The volatile organic compounds benzene, toluene and xylenes (BTX) are among the most produced chemicals worldwide and can be found in different environmental compartments. Contamination of aquatic environments by these substances may result in to adverse toxic effects on organisms of different biological organizations. The present study aimed to evaluate the toxicity of BTX isolated and in binary mixtures using the acute toxicity test (96h) with the microcrustacean Mysidopsis juniae. To achieve that, organisms were exposed to BTX concentrations in order to identify the effects on survival, as well as to study the modes of action of the substances in mixture with the application of the theoretical models of Addition Concentration (AC) and Independent Action (AI). In the experiments with the isolated compounds it was possible to notice that xylene was the most toxic compound (LC50 16,10 ±2,4mg.L-1) followed by toluene (LC50 38 ± 5,3mg.L-1) and benzene (LC50 78,03 ±2,9mg.L-1) respectively. When in binary mixture deviations of the reference models for CA were observed in all experiments performed (dose ratio and antagonism). In the exposure of xylene and benzene the deviation that fit the results was the antagonism, where as the mixture of xylene and toluene was better explained by the deviation dose ratio where the toxicity was caused mainly by xylene. In the experiment combining toluene and benzene, the dose ratio pattern was observed as well, and the benzene was the main responsible for the toxicity. We concluded that BTX were toxic to mysids, both isolated and in binary mixtures, evidencing the importance of studies in this subject that can serve as support for evaluations and monitoring of the marine environments, considering the imminent risks of contamination of this environment and the wide scale of use of these compounds. / Os compostos orgânicos voláteis benzeno, tolueno e os xilenos (BTX) estão entre os produtos químicos mais produzidos mundialmente e podem ser encontrados em diferentes compartimentos ambientais. A contaminação de ambientes aquáticos por essas substâncias pode acarretar em efeitos tóxicos adversos em organismos de diferentes organizações biológicas. Diante disso, o presente estudo teve como objetivo avaliar a toxicidade dos BTX isolados e em misturas binárias por meio do teste de toxicidade aguda (96h) com o microcrustáceo Mysidopsis juniae. Para isso, os organismos foram expostos às concentrações de BTX a fim de identificar os efeitos na sobrevivência, assim como estudar os modos de ação das substâncias em mistura com a aplicação dos modelos teóricos de concentração de adição (CA) e ação independente (IA). Nos experimentos com os compostos isolados foi possível perceber que o xileno foi o mais tóxico (CL50 16,10±2,4mg.L-1) seguido por tolueno (CL50 38±5,3mg.L-1) e benzeno (CL50 78,03±2,9mg.L-1). Quando em mistura binária foram observados desvios dos modelos de referência para CA em todos os experimentos realizados (razão das doses e antagonismo). Na exposição do xileno e benzeno o desvio que se enquadrou aos resultados foi o antagonismo, já a mistura de xileno e tolueno foi melhor explicada pelo desvio razão das doses onde a toxicidade foi causada principalmente pelo xileno, no experimento com o tolueno e benzeno também foi observado um padrão de razão das doses, sendo que nesse caso o benzeno foi o maior responsável pela toxicidade. Logo, os hidrocarbonetos analisados foram tóxicos para o misidáceo, tanto isolados como em misturas binárias, evidenciando a importância de estudos nessa temática que possam servir de suporte para avaliações e monitoramento dos ambientes marinhos, visto os riscos iminentes de contaminação deste meio e a ampla escala de utilização desses compostos.

En av de vanligaste föroreningarna i mark och grundvatten är aromatiska kolvä- ten såsom bensen, toluen, etylbensen och xylener. Dessa härstammar från ex. ben- sin och har läckt ut till omgivande miljö från drivmedelsanläggningar. I stället för schaktning och bortforsling av de förorenade jordmassorna som mest förekom- mande ex situ-saneringsmetod behövs mer forskning kring in situ-saneringsmetoder där föroreningen alltså kan angripas på plats. I denna rapport undersöks förhöjd biologisk nedbrytning genom att redogöra för de önskvärda (bio)kemiska reak- tionerna i marken, de parametrar som styr den biologiska nedbrytningen samt huruvida dessa parametrars inverkan har kunnat påvisas i tre stycken fallstu- dier där in situ-sanering utförts vid drivmedelsanläggningar. Studien kommer fram till ett antal primära (ursprungliga) variabler såsom föroreningen i fråga, jordtätheten och klimatet, samt ett antal sekundära (förändringsbara) variabler såsom oxidationsmedlet, mikroorganismerna, näringsämnena och markvattnet. Dessa presenterades sedan i en schematisk skiss som visar hur de växelverkar med parametrarna för optimal biologisk nedbrytning, vilka visade sig ha vissa optimala värden. Dessa konstaterades vara föroreningshalten (<5-10 viktprocent torr jord), pH-värdet (6-9), temperaturen (20-35◦C), vattenmättnaden (40-80%), syrehalten (> 2 500 viktprocent kolväten), C:N:P-kvoten (100:10:1) samt koncent- rationen mikroorganismer (> 103 CFU/ml). När de olika parametrarnas inverkan  i fallstudierna undersöktes kunde slutsatsen dras att det skulle behövas ytterliga- re data och mätningar vid varje sanering för att kunna garantera att parametrarna varit inom sina respektive optimala värden. / One of the many goals for a sustainable living and a healthy environment is the reduction of toxic substances in soil and groundwater. Within this ambit, one of the most common pollutants is aromatic hydrocarbons such as benzene, tolue- ne, ethylbenzene and xylenes (BTEX) which are spread from e.g. filling stations for fuel as they are constituents of petrol. These are volatile and can constitute a risk for humans and the environment as they percolate through the soil and groundwater resources as well as can exfiltrate from the ground to surrounding buildings and pollute the indoor air. Once these pollutants are present in the soil, the site needs to be remediated, something which is mainly done ex situ by exca- vating the polluted masses to be sent to a treatment facility. An alternative to this is in situ remediation where the pollution can be treated without being excavated and thus having a smaller impact on the environment.  In this report, one of these techniques called enhanced biodegradation was studied, which implies the application of oxidisers to faciliate microbial activities. It investigated (1) how this remediation technique works and what (bio)chemical reactions are desired in the soil; (2) what are the principal parameters that enable this remediation process and how do they interact with the surrounding factors, and (3) whether the impact of these parameters can be estimated in three case studies where this technique has been used. The three case studies were filling stations located in Sweden, having similar pollutants and geology, and two diffe- rent oxidisers had been used.  The study resulted in the identification of a number of primary (initial) varia- bles such as the pollutant characteristics, the soil density and the climate; as well as a number of secodnary (modifiable) variables such as the oxidiser, microorga- nisms, nutrients and soil moisture. These were then presented in a scheme which shows how their interactions with the parameters for optimal biodegradation. Each parameter was found to have a certain optimum such as the concentration of hydrocarbons (<5-10 mass percent of the dry soil) , pH value (6-9), tempe- rature (20-35◦C), soil water saturation (40-80%), oxygen concentration (> 2,500 mass percent of the hydrocarbons), C:N:P ratio (100:10:1) as well as concentration of microorganisms (> 103 CFU/ml). A negative correlation could be identified between the soil density and the oxygen concentration in the soil, as well as a po- sitive correlation between the soil water saturation in the and the soil moisture, alternatively additional water via irrigation. The impact of each parameter in the case studies of in situ remediation was assessed and the it could be concluded that they would need additional data and measurements for most parameters in order to guarantee that they are within the respective optima.