The purpose of this study was to determine the chemical and/or biological factors that cause 2,4-dinitrotoluene (2,4-DNT), 2,6-dinitrotoluene (2,6-DNT) and nitrobenzene (NB) to transform to their respective aromatic amines in the Borden aquifer, and to investigate the biodegradation of 2,4-diaminotoluene (2,4-DAT) and 2,6-diaminoluene (2,6-DAT) under aerobic conditions. In situ microcosms (ISM) and laboratory microcosm experiments were used in the investigation. In addition, a sequential treatment system was tested in which columns containing granular iron were followed by either an anaerobic or aerobic soil column. Both 2,4- and 2,6-DNT were used to determine if competitive effects exist between the two. The ISM isolates a volume of the aquifer material and allows for in situ solute loading and sampling in order to characterize chemical or biological reactions. Four ISMs were installed below the water table at CFB Borden. Each ISM was injected with 10 mg/L of either 2,4-DNT, 2,6-DNT, NB, or 2,4-DNT + 2,6-DNT, in two repetitions. In all cases, chloride was also injected as a conservative tracer to monitor for dilution. The results indicated transformation of nitroaromatics via nitro-reduction to their intermediate products, mainly as 2,4-DAT, 2,6-DAT, and aniline. Within 20 days, a loss of up to 92% of 2,4-DNT was observed with the formation of 2,4-DAT. Minor amounts of 2-amino-4-nitrotoluene (2-A-4-NT) and 4-amino-2-nitrotoluene (4-A-2-NT) were also observed. Similarly, up to a 96% loss of 2,6-DNT was seen after 29 days, with degradation products including 2-amino-6-nitrotoluene (2-A-6-NT) and 2,6-DAT. When 2,4- and 2,6-DNT were present in combination, 99% loss of both compounds at similar rates was observed over 20 days following the injections, with degradation products including aminonitrotoluenes and diaminotoluenes. Finally, when nitrobenzene was injected, degradation of up to 99% was observed by day 29, with the formation of aniline as the primary product. To determine the cause of the nitro-reduction, laboratory microcosm experiments were conducted using soil from within the chamber of the ISM’s. Duplicate microcosms were prepared with Borden groundwater and spiked with 2,4- and 2,6-DNT in an anaerobic glovebox. Microcosms were incubated and sampled periodically for approximately 3 months. Several different conditions, including: groundwater and soil, autoclaved groundwater and soil, soil taken at ground surface and groundwater, and autoclaved silica sand and groundwater were created for microcosm experiments to determine whether abiotic or biotic factors caused the reduction of 2,4- and 2,6-DNT. Microcosms which duplicated field conditions in the laboratory had average half-lives of 4.2 days and 5.1 days for 2,4- and 2,6-DNT, respectively, compared to the field result with average half-lives between 3.9 days (2,4-DNT) and 3.5 days (2,6-DNT). Subsequently, a nutrient medium was added to each repetition. The behaviour of DNT degradation did not change significantly, suggesting minimal involvement of biological processes. Furthermore soil analysis showed relatively high concentrations of extractable iron and the presence of magnetite, which are species capable of reducing nitroaromatics. Therefore, it is concluded that nitro-reduction in Borden soil is likely a result of abiotic surface mediated processes. The competitive behaviour of 2,4- and 2,6-DNT was studied in a sequential treatment system which consisted of an anaerobic iron column, followed by either an anaerobic or aerobic soil column. Results showed the same rate of transformation from 2,4- and 2,6-DNT within the iron column, with 100% conversion to 2,4- and 2,6-DAT, respectively. Within the anaerobic and aerobic soil columns, the DATs were highly persistent. When a nutrient solution was added only to the aerobic soil column with DNTs as the initial compounds, results showed a reduction of 2,4-DNT of 17%, with an increase in 2,6-DNT of 22%. The increase in 2,6-DNT may have been a result of differing influent concentrations at earlier pore volumes. When stock solutions in the aerobic column were altered to only include DATs, a reduction of 2,4- and 2,6-DAT was observed at 17% and 18%, respectively. It would appear that an acclimated bacterial community able to transform DNT and DAT was present in the aerobic Borden sand column. Degradation of 2,4- and 2,6-DAT was dependant on the degree of nutrients supplied to indigenous bacterial communities under aerobic conditions.

The purpose of this study was to determine the chemical and/or biological factors that cause 2,4-dinitrotoluene (2,4-DNT), 2,6-dinitrotoluene (2,6-DNT) and nitrobenzene (NB) to transform to their respective aromatic amines in the Borden aquifer, and to investigate the biodegradation of 2,4-diaminotoluene (2,4-DAT) and 2,6-diaminoluene (2,6-DAT) under aerobic conditions. In situ microcosms (ISM) and laboratory microcosm experiments were used in the investigation. In addition, a sequential treatment system was tested in which columns containing granular iron were followed by either an anaerobic or aerobic soil column. Both 2,4- and 2,6-DNT were used to determine if competitive effects exist between the two. The ISM isolates a volume of the aquifer material and allows for in situ solute loading and sampling in order to characterize chemical or biological reactions. Four ISMs were installed below the water table at CFB Borden. Each ISM was injected with 10 mg/L of either 2,4-DNT, 2,6-DNT, NB, or 2,4-DNT + 2,6-DNT, in two repetitions. In all cases, chloride was also injected as a conservative tracer to monitor for dilution. The results indicated transformation of nitroaromatics via nitro-reduction to their intermediate products, mainly as 2,4-DAT, 2,6-DAT, and aniline. Within 20 days, a loss of up to 92% of 2,4-DNT was observed with the formation of 2,4-DAT. Minor amounts of 2-amino-4-nitrotoluene (2-A-4-NT) and 4-amino-2-nitrotoluene (4-A-2-NT) were also observed. Similarly, up to a 96% loss of 2,6-DNT was seen after 29 days, with degradation products including 2-amino-6-nitrotoluene (2-A-6-NT) and 2,6-DAT. When 2,4- and 2,6-DNT were present in combination, 99% loss of both compounds at similar rates was observed over 20 days following the injections, with degradation products including aminonitrotoluenes and diaminotoluenes. Finally, when nitrobenzene was injected, degradation of up to 99% was observed by day 29, with the formation of aniline as the primary product. To determine the cause of the nitro-reduction, laboratory microcosm experiments were conducted using soil from within the chamber of the ISM’s. Duplicate microcosms were prepared with Borden groundwater and spiked with 2,4- and 2,6-DNT in an anaerobic glovebox. Microcosms were incubated and sampled periodically for approximately 3 months. Several different conditions, including: groundwater and soil, autoclaved groundwater and soil, soil taken at ground surface and groundwater, and autoclaved silica sand and groundwater were created for microcosm experiments to determine whether abiotic or biotic factors caused the reduction of 2,4- and 2,6-DNT. Microcosms which duplicated field conditions in the laboratory had average half-lives of 4.2 days and 5.1 days for 2,4- and 2,6-DNT, respectively, compared to the field result with average half-lives between 3.9 days (2,4-DNT) and 3.5 days (2,6-DNT). Subsequently, a nutrient medium was added to each repetition. The behaviour of DNT degradation did not change significantly, suggesting minimal involvement of biological processes. Furthermore soil analysis showed relatively high concentrations of extractable iron and the presence of magnetite, which are species capable of reducing nitroaromatics. Therefore, it is concluded that nitro-reduction in Borden soil is likely a result of abiotic surface mediated processes. The competitive behaviour of 2,4- and 2,6-DNT was studied in a sequential treatment system which consisted of an anaerobic iron column, followed by either an anaerobic or aerobic soil column. Results showed the same rate of transformation from 2,4- and 2,6-DNT within the iron column, with 100% conversion to 2,4- and 2,6-DAT, respectively. Within the anaerobic and aerobic soil columns, the DATs were highly persistent. When a nutrient solution was added only to the aerobic soil column with DNTs as the initial compounds, results showed a reduction of 2,4-DNT of 17%, with an increase in 2,6-DNT of 22%. The increase in 2,6-DNT may have been a result of differing influent concentrations at earlier pore volumes. When stock solutions in the aerobic column were altered to only include DATs, a reduction of 2,4- and 2,6-DAT was observed at 17% and 18%, respectively. It would appear that an acclimated bacterial community able to transform DNT and DAT was present in the aerobic Borden sand column. Degradation of 2,4- and 2,6-DAT was dependant on the degree of nutrients supplied to indigenous bacterial communities under aerobic conditions.

A biomassa lignocelulósica, como o bagaço de cana-de-açúcar, tem potencial para ser usado como matéria-prima na fabricação de produtos de valor agregado, uma vez que, seus componentes estruturais podem ser separados através de pré-tratamentos e utilizados em linhas de processos. Diferentes tipos de pré-tratamentos tem sido desenvolvidos com este objetivo, e neste contexto, foi proposto um tratamento sequencial químico-enzimático (SQE) do bagaço de cana-de-açúcar com três estágios; 1) Extração alcalina a frio (CAE): realizado com 10% (m/m) de NaOH por 30 min a 25ºC, 2) Pré-tratamento sulfito alcalino em etanol (ASE): realizado com 2,5% (m/m) de NaOH e 5% (m/m) de Na2SO3 em etanol (30 %v/v), por 2 h a 120ºC e 3) Extração enzimática da hemicelulose residual (EEH): conduzida com extrato comercial de xilanase (Luminase) a 5UI/g de biomassa em tampão fosfato de sódio 50 mM, pH 8 a 50ºC, por 6 horas e 24 horas. O tratamento SQE permitiu a solubilização de 48% e 60% da hemicelulose e 86% e 84% da lignina original do bagaço, diferenças obtidas em função do tempo de extração enzimática de 6 e 24 horas, respectivamente. Os sólidos resultantes da segunda etapa do pré-tratamento (polpa-P2) e da terceira etapa (polpa-P3) foram hidrolisados com o coquetel enzimático Cellic Ctec2 (10 FPU/g de glucana) por 48h a 50ºC, pH 4,8, nas consistências de 5%, 10% e 15% m/v. A extração enzimática de hemiceluloses (terceira etapa do tratamento SQE) da polpa-P2 não contribuiu com a hidrólise de celulose. Na consistência de 5%, as polpas P2 e P3 apresentaram 95 e 94% de conversão de celulose em 24h, valores similares foram obtidos para as polpas na consistência de 10%, porém em 48h de reação. A conversão de celulose das polpas P2 e P3 em 48h, a 15% de consistência, diminuiu para 84% e 81%, respectivamente. A polpa P3, proveniente da extração enzimática das hemiceluloses por 24h, apresentou um menor valor de conversão de celulose (74%), a 15% de consistência, evidenciando-se o efeito negativo da extração adicional de hemicelulose sobre a hidrólise da celulose. Embora não tenham sido observadas diferenças significativas nas porcentagens de conversão de celulose nas polpas P2 e P3, a implementação das três etapas de pré-tratamentos possibilitou a obtenção de duas frações diferentes de hemiceluloses, que foram recuperadas por precipitação com etanol, cada uma delas com características e aplicações potenciais diferentes. A composição química das hemiceluloses extraídas do bagaço de cana as define como arabinoxilana. As condições operacionais utilizadas na primeira etapa (CAE) do tratamento SQE gerou xilanas com maiores massas molares (34.598 g/mol) e mais contaminadas com lignina (18%) comparadas às xilanas recuperadas na terceira etapa (EEH), que apresentaram massas molares entre 9.948-11.678g/mol com 1,5- 3,5% de lignina. Nestas últimas foram identificados a presença de xilooligossacarideos (XOS) como xilotriose (X3), xilotetraose (X4) e xilopentaose (X5). / Lignocellulosic biomass such as sugarcane bagasse has the potential to be used as raw material in the manufacture of value-added products, since its structural components can be separated through pre-treatments and used in process lines. Different types of pretreatments have been developed with this objective, and in this context, a sequential chemical-enzymatic treatment (SQE) of three-stage sugarcane bagasse was proposed. 1) Cold alkaline extraction (CAE): performed with 10% (w/w) NaOH for 30 min at 25ºC, 2) Alkaline sulfite etanol pre-treatment (ASE): performed with 2.5% (w/w) NaOH and 5% (w/w) Na2SO3 in ethanol (30% v/v) for 2h at 120ºC and 3) Enzymatic extraction of residual hemicellulose (EEH): Conducted with commercial extract of xylanase (Luminase) at 5UI/g biomass in 50mM sodium phosphate buffer, pH 8 at 50ºC, for 6h and 24h. The SQE treatment allowed the solubilization of 48% and 60% of the hemicellulose and 86% and 84% of the original bagasse lignin, differences obtained as a function of the enzymatic extraction time of 6 and 24 hours, respectively. The solids resulting from the second stage (pulp P2) and the third stage (pulp P3) of the pretreatment were hydrolyzed with the enzymatic cocktail Cellic Ctec2 (10FPU/g glucan) for 48h at 50ºC pH 4.8, in the consistencies of 5%, 10% and 15% m/v. The enzymatic extraction of hemicelluloses (third stage of the treatment SQA) of the pulp-P2 did not contribute to the hydrolysis of cellulose. At the consistency of 5%, pulps P2 and P3 presented 95 and 94% of cellulose conversion in 24h, similar values were obtained for those pulps in the consistency of 10%, but in 48h of reaction. The cellulose conversion of pulps P2 and P3 in 48h, at 15% consistency decreased to 84% and 81%, respectively. The pulp P3, from the enzymatic extraction of the hemicelluloses for 24h, presented a lower value of cellulose conversion (74%), at 15% of consistency, evidencing the negative effect of the additional extraction of hemicellulose on the hydrolysis of cellulose. Although no significant differences were observed in the cellulose conversion percentages in the P2 and P3 pulps, the implementation of the three pretreatment steps allowed two different fractions of hemicelluloses to be obtained, which were recovered by precipitation with ethanol, each with characteristics and potential applications. The chemical composition of the hemicelluloses extracted from the sugarcane bagasse describes them as arabinoxylan. The operating conditions used in the first stage (CAE) of the SQE treatment generated xylans with higher molar masses (34,598 g/mol) and more lignin contaminants (18%) compared to the third stage (EEH) recovered xylans, which presented molar masses between 9,948-11,678g/mol with 1.5-3.5% lignin. In the latter, the presence of xylo-oligosaccharides (XOS) such as xylotriose (X3), xylotetraose (X4) and xylopentaose (X5) were identified.

A biomassa lignocelulósica, como o bagaço de cana-de-açúcar, tem potencial para ser usado como matéria-prima na fabricação de produtos de valor agregado, uma vez que, seus componentes estruturais podem ser separados através de pré-tratamentos e utilizados em linhas de processos. Diferentes tipos de pré-tratamentos tem sido desenvolvidos com este objetivo, e neste contexto, foi proposto um tratamento sequencial químico-enzimático (SQE) do bagaço de cana-de-açúcar com três estágios; 1) Extração alcalina a frio (CAE): realizado com 10% (m/m) de NaOH por 30 min a 25ºC, 2) Pré-tratamento sulfito alcalino em etanol (ASE): realizado com 2,5% (m/m) de NaOH e 5% (m/m) de Na2SO3 em etanol (30 %v/v), por 2 h a 120ºC e 3) Extração enzimática da hemicelulose residual (EEH): conduzida com extrato comercial de xilanase (Luminase) a 5UI/g de biomassa em tampão fosfato de sódio 50 mM, pH 8 a 50ºC, por 6 horas e 24 horas. O tratamento SQE permitiu a solubilização de 48% e 60% da hemicelulose e 86% e 84% da lignina original do bagaço, diferenças obtidas em função do tempo de extração enzimática de 6 e 24 horas, respectivamente. Os sólidos resultantes da segunda etapa do pré-tratamento (polpa-P2) e da terceira etapa (polpa-P3) foram hidrolisados com o coquetel enzimático Cellic Ctec2 (10 FPU/g de glucana) por 48h a 50ºC, pH 4,8, nas consistências de 5%, 10% e 15% m/v. A extração enzimática de hemiceluloses (terceira etapa do tratamento SQE) da polpa-P2 não contribuiu com a hidrólise de celulose. Na consistência de 5%, as polpas P2 e P3 apresentaram 95 e 94% de conversão de celulose em 24h, valores similares foram obtidos para as polpas na consistência de 10%, porém em 48h de reação. A conversão de celulose das polpas P2 e P3 em 48h, a 15% de consistência, diminuiu para 84% e 81%, respectivamente. A polpa P3, proveniente da extração enzimática das hemiceluloses por 24h, apresentou um menor valor de conversão de celulose (74%), a 15% de consistência, evidenciando-se o efeito negativo da extração adicional de hemicelulose sobre a hidrólise da celulose. Embora não tenham sido observadas diferenças significativas nas porcentagens de conversão de celulose nas polpas P2 e P3, a implementação das três etapas de pré-tratamentos possibilitou a obtenção de duas frações diferentes de hemiceluloses, que foram recuperadas por precipitação com etanol, cada uma delas com características e aplicações potenciais diferentes. A composição química das hemiceluloses extraídas do bagaço de cana as define como arabinoxilana. As condições operacionais utilizadas na primeira etapa (CAE) do tratamento SQE gerou xilanas com maiores massas molares (34.598 g/mol) e mais contaminadas com lignina (18%) comparadas às xilanas recuperadas na terceira etapa (EEH), que apresentaram massas molares entre 9.948-11.678g/mol com 1,5- 3,5% de lignina. Nestas últimas foram identificados a presença de xilooligossacarideos (XOS) como xilotriose (X3), xilotetraose (X4) e xilopentaose (X5). / Lignocellulosic biomass such as sugarcane bagasse has the potential to be used as raw material in the manufacture of value-added products, since its structural components can be separated through pre-treatments and used in process lines. Different types of pretreatments have been developed with this objective, and in this context, a sequential chemical-enzymatic treatment (SQE) of three-stage sugarcane bagasse was proposed. 1) Cold alkaline extraction (CAE): performed with 10% (w/w) NaOH for 30 min at 25ºC, 2) Alkaline sulfite etanol pre-treatment (ASE): performed with 2.5% (w/w) NaOH and 5% (w/w) Na2SO3 in ethanol (30% v/v) for 2h at 120ºC and 3) Enzymatic extraction of residual hemicellulose (EEH): Conducted with commercial extract of xylanase (Luminase) at 5UI/g biomass in 50mM sodium phosphate buffer, pH 8 at 50ºC, for 6h and 24h. The SQE treatment allowed the solubilization of 48% and 60% of the hemicellulose and 86% and 84% of the original bagasse lignin, differences obtained as a function of the enzymatic extraction time of 6 and 24 hours, respectively. The solids resulting from the second stage (pulp P2) and the third stage (pulp P3) of the pretreatment were hydrolyzed with the enzymatic cocktail Cellic Ctec2 (10FPU/g glucan) for 48h at 50ºC pH 4.8, in the consistencies of 5%, 10% and 15% m/v. The enzymatic extraction of hemicelluloses (third stage of the treatment SQA) of the pulp-P2 did not contribute to the hydrolysis of cellulose. At the consistency of 5%, pulps P2 and P3 presented 95 and 94% of cellulose conversion in 24h, similar values were obtained for those pulps in the consistency of 10%, but in 48h of reaction. The cellulose conversion of pulps P2 and P3 in 48h, at 15% consistency decreased to 84% and 81%, respectively. The pulp P3, from the enzymatic extraction of the hemicelluloses for 24h, presented a lower value of cellulose conversion (74%), at 15% of consistency, evidencing the negative effect of the additional extraction of hemicellulose on the hydrolysis of cellulose. Although no significant differences were observed in the cellulose conversion percentages in the P2 and P3 pulps, the implementation of the three pretreatment steps allowed two different fractions of hemicelluloses to be obtained, which were recovered by precipitation with ethanol, each with characteristics and potential applications. The chemical composition of the hemicelluloses extracted from the sugarcane bagasse describes them as arabinoxylan. The operating conditions used in the first stage (CAE) of the SQE treatment generated xylans with higher molar masses (34,598 g/mol) and more lignin contaminants (18%) compared to the third stage (EEH) recovered xylans, which presented molar masses between 9,948-11,678g/mol with 1.5-3.5% lignin. In the latter, the presence of xylo-oligosaccharides (XOS) such as xylotriose (X3), xylotetraose (X4) and xylopentaose (X5) were identified.