The biotechnology of hard coal utilization as a bioprocess substrate
- Mutambanengwe, Cecil Clifford Zvandada
- Authors: Mutambanengwe, Cecil Clifford Zvandada
- Date: 2010
- Subjects: Coal -- Biotechnology Acid mine drainage Coal mines and mining -- Environmental aspects
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3934 , http://hdl.handle.net/10962/d1003993
- Description: The development of coal biotechnology, using hard coal as a substrate, has been impeded by its low reactivity in biological processes. As a result, the more successful application studies have focused on lignitic soft coals. However, new studies have reported using biologically or geologically oxidized hard coal as a functional substrate option for bioprocess applications on a large scale. This study undertook a preliminary investigation into the feasibility of environmental applications of coal biotechnology using oxidized hard coal substrates in both anaerobic and aerobic processes with carbon dioxide, sulfate and oxygen as terminal electron acceptors. A preliminary characterization of the oxidized hard coal substrates was undertaken to determine and predict their viability and behavior as electron donors and carbon sources for environmental bioprocess applications of direct interest to the coal mining industry. Both biologically and geologically oxidized coal substrates showed loss of up to 17% and 52% carbon respectively and incorporation of oxygen ranging from 0.9 – 24%. The latter substrate showed greater loss of carbon and increased oxygenation. The biologically and geologically oxidized hard coal substrates were shown to partition readily into 23% and 32% organic humic acid, a 0.1% fulvic acid fraction and 65% and 59% inorganic and humin fractions respectively. These organic components were shown to be potentially available for biological consumption. In the unmodified hard coal substrate, partitioning was not observed and it did not perform as a functional substrate for any of the bioprocesses investigated. Where carbon dioxide was used as a terminal electron acceptor, methane production ranging from 9 – 26 mg CH4.g substrate-1 was demonstrated from both oxidized coal substrates. Geologically oxidized coal produced 30% more methane than biologically oxidized coal. Methane yields from the geologically oxidized coal in the presence and absence of a co-substrate were 5 – 13-fold higher than previous studies that used hard coal for methanogenesis. Based on these results, and that the development and optimization of the biological oxidation process is currently ongoing, further applications investigated in this study were undertaken using geologically oxidized coal. It was shown using pyrolysis gas chromatography mass spectrometry that the methanogenic system was dependent on the presence of an effective co-substrate supporting the breakdown of the complex organic structures within the oxidized hard coal substrate. Also that the accumulation of aromatic intermediate breakdown compounds predominantly including toluene, furfural, styrene and 2-methoxy vinyl phenol appeared to become inhibitory to both methanogenic and sulfidogenic reactions. This was shown to be a more likely cause of reactor failure rather than substrate exhaustion over time. Evidence of a reductive degradation pathway of the complex organic structures within the oxidized hard coal substrates was shown through the production, accumulation and utilization of volatile fatty acids including acetic, formic, propionic, butyric and valeric acids. Comparative analysis of the volatile fatty acids produced in this system showed that geologically oxidized coal produced 20% more of the volatile fatty acids profiled and double the total concentration compared to the biologically oxidized coal. The use of geologically oxidized hard coal as a functional substrate for biological sulfate reduction was demonstrated in the neutralization of a simulated acid mine drainage wastewater in both batch and continuous process operations. Results showed an increase in pH from pH 4.0 to ~ pH 8.0 with sulfide production rates of ~ 86 mgL-1.day-1 in the batch reactions, while the pH increased to pH 9.0 and sulfide production rates of up to 450 mgL-1.day-1 were measured in the continuous process studies using sand and coal up-flow packed bed reactors. Again, the requirement for an effective co-substrate was demonstrated with lactate shown to function as a true co-substrate in this system. However, a low cost alternative to lactate would need to emerge if the process was to function in large-scale commercial environmental treatment applications. In this regard, the aerobic growth and production of Neosartorya fischeri biomass (0.64 g.biomass.g SOC-1) was demonstrated using oxidized hard coal and glutamate as a co-substrate. Both can be produced from wastes generated on coal mines, with the fungal biomass generated in potentially large volumes. Preliminary demonstration of the use of the fungal biomass as a carbon and electron donor source for biological sulfate reduction was shown and thus that this could serve as an effective substrate for anaerobic environmental treatment processes. Based on these findings, an Integrated Coal Bioprocess model was proposed using oxidized hard coal as a substrate for environmental remediation applications on coal mines. In this approach, potential applications included methane recovery from waste coal, use of waste coal in the treatment of acid mine drainage waste waters and the recovery and use of humic acids in the rehabilitation of open cast mining soils. This study provided a first report demonstrating the use of biologically and geologically oxidized hard coals as bioprocess substrates in environmental bioremediation applications. It also provided an indication that follow-up bioengineering studies to investigate scaled-up applications of these findings would be warranted.
- Full Text:
- Authors: Mutambanengwe, Cecil Clifford Zvandada
- Date: 2010
- Subjects: Coal -- Biotechnology Acid mine drainage Coal mines and mining -- Environmental aspects
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:3934 , http://hdl.handle.net/10962/d1003993
- Description: The development of coal biotechnology, using hard coal as a substrate, has been impeded by its low reactivity in biological processes. As a result, the more successful application studies have focused on lignitic soft coals. However, new studies have reported using biologically or geologically oxidized hard coal as a functional substrate option for bioprocess applications on a large scale. This study undertook a preliminary investigation into the feasibility of environmental applications of coal biotechnology using oxidized hard coal substrates in both anaerobic and aerobic processes with carbon dioxide, sulfate and oxygen as terminal electron acceptors. A preliminary characterization of the oxidized hard coal substrates was undertaken to determine and predict their viability and behavior as electron donors and carbon sources for environmental bioprocess applications of direct interest to the coal mining industry. Both biologically and geologically oxidized coal substrates showed loss of up to 17% and 52% carbon respectively and incorporation of oxygen ranging from 0.9 – 24%. The latter substrate showed greater loss of carbon and increased oxygenation. The biologically and geologically oxidized hard coal substrates were shown to partition readily into 23% and 32% organic humic acid, a 0.1% fulvic acid fraction and 65% and 59% inorganic and humin fractions respectively. These organic components were shown to be potentially available for biological consumption. In the unmodified hard coal substrate, partitioning was not observed and it did not perform as a functional substrate for any of the bioprocesses investigated. Where carbon dioxide was used as a terminal electron acceptor, methane production ranging from 9 – 26 mg CH4.g substrate-1 was demonstrated from both oxidized coal substrates. Geologically oxidized coal produced 30% more methane than biologically oxidized coal. Methane yields from the geologically oxidized coal in the presence and absence of a co-substrate were 5 – 13-fold higher than previous studies that used hard coal for methanogenesis. Based on these results, and that the development and optimization of the biological oxidation process is currently ongoing, further applications investigated in this study were undertaken using geologically oxidized coal. It was shown using pyrolysis gas chromatography mass spectrometry that the methanogenic system was dependent on the presence of an effective co-substrate supporting the breakdown of the complex organic structures within the oxidized hard coal substrate. Also that the accumulation of aromatic intermediate breakdown compounds predominantly including toluene, furfural, styrene and 2-methoxy vinyl phenol appeared to become inhibitory to both methanogenic and sulfidogenic reactions. This was shown to be a more likely cause of reactor failure rather than substrate exhaustion over time. Evidence of a reductive degradation pathway of the complex organic structures within the oxidized hard coal substrates was shown through the production, accumulation and utilization of volatile fatty acids including acetic, formic, propionic, butyric and valeric acids. Comparative analysis of the volatile fatty acids produced in this system showed that geologically oxidized coal produced 20% more of the volatile fatty acids profiled and double the total concentration compared to the biologically oxidized coal. The use of geologically oxidized hard coal as a functional substrate for biological sulfate reduction was demonstrated in the neutralization of a simulated acid mine drainage wastewater in both batch and continuous process operations. Results showed an increase in pH from pH 4.0 to ~ pH 8.0 with sulfide production rates of ~ 86 mgL-1.day-1 in the batch reactions, while the pH increased to pH 9.0 and sulfide production rates of up to 450 mgL-1.day-1 were measured in the continuous process studies using sand and coal up-flow packed bed reactors. Again, the requirement for an effective co-substrate was demonstrated with lactate shown to function as a true co-substrate in this system. However, a low cost alternative to lactate would need to emerge if the process was to function in large-scale commercial environmental treatment applications. In this regard, the aerobic growth and production of Neosartorya fischeri biomass (0.64 g.biomass.g SOC-1) was demonstrated using oxidized hard coal and glutamate as a co-substrate. Both can be produced from wastes generated on coal mines, with the fungal biomass generated in potentially large volumes. Preliminary demonstration of the use of the fungal biomass as a carbon and electron donor source for biological sulfate reduction was shown and thus that this could serve as an effective substrate for anaerobic environmental treatment processes. Based on these findings, an Integrated Coal Bioprocess model was proposed using oxidized hard coal as a substrate for environmental remediation applications on coal mines. In this approach, potential applications included methane recovery from waste coal, use of waste coal in the treatment of acid mine drainage waste waters and the recovery and use of humic acids in the rehabilitation of open cast mining soils. This study provided a first report demonstrating the use of biologically and geologically oxidized hard coals as bioprocess substrates in environmental bioremediation applications. It also provided an indication that follow-up bioengineering studies to investigate scaled-up applications of these findings would be warranted.
- Full Text:
Hydrogenases from sulphate reducing bacteria and their role in the bioremediation of textile effluent
- Mutambanengwe, Cecil Clifford Zvandada
- Authors: Mutambanengwe, Cecil Clifford Zvandada
- Date: 2007
- Subjects: Bioremediation , Dyes and dyeing -- Waste disposal , Sulfur bacteria , Hydragenase , Factory and trade waste -- Purification , Textile waste
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3960 , http://hdl.handle.net/10962/d1004019 , Bioremediation , Dyes and dyeing -- Waste disposal , Sulfur bacteria , Hydragenase , Factory and trade waste -- Purification , Textile waste
- Description: The continuing industrial development has led to a corresponding increase in the amount of waste water generation leading to a consequential decline in levels and quality of the natural water in the ecosystem. Textile industries consume over 7 x 10[superscript 5] tons of dyes annually and use up to 1 litre of water per kg of dye processed and are third largest polluters in the world, the problem being aggravated by the inefficiencies of the dye houses. An abundance of physio-chemical methods are in use world wide, however, there is increasing concern as to their impact in effectively treating textile effluents as they introduce secondary pollutants during the ‘remediation’ process which are quite costly to run, maintain and clean up. Research on biological treatment has offered simple and cost effective ways of bioremediating textile effluents. While aerobic treatment of textile dyes and their effluents has been reported, its major draw back is commercial up-scaling and as such anaerobic systems have been investigated and shown to degrade azo dyes, which form the bulk of the dyes used world wide. However, the mechanisms involved in the bioremediation of these dyes are poorly understood. The aims of this study were to identify and investigate the role of enzymes produced by sulphate reducing bacteria (SRB) in bioremediating textile dye and their effluents. Sulphate reducing bacteria were used in this study because they are tolerant to harsh environmental conditions and inhibit the proliferance of pathogenic micro-organisms. The appearance of clear zones in agar plates containing azo dye concentrations ranging from 10 – 100 mgl[superscript -1] showed the ability of SRB to decolourize dyes under anaerobic conditions. Assays of enzymes previously reported to decolourise azo dyes were not successful, but led to the identification of hydrogenase enzyme being produced by SRB. The enzyme was found to be localised in the membrane and cytoplasm. A surface response method was used to optimize the extraction of the enzyme from the bacterial cells resulting in approximately 3 fold increase in hydrogenase activity. Maximum hydrogenase activity was found to occur after six days in the absence of dyes but was found to occur after one day in the presence of azo dyes. A decline in hydrogenase activity thereafter, suggested inhibition of enzymatic activity by the putative aromatic amines produced after azo cleavage. Purification of the hydrogenase by freeze drying, poly ethylene glycol, and Sephacryl – 200 size exclusion- ion exchange chromatography revealed the enzyme to have a molecular weight of 38.5 kDa when analyzed by a 12 % SDS-PAGE. Characterisation of the enzyme revealed optimal activity at a pH of 7.5 and temperature of 40 °C while it exhibited a poor thermal stability with a half-life of 32 minutes. The kinetic parameters V[subscript max] and K[subscript m] were 21.18 U ml[superscript -1} and 4.57 mM respectively. Application of the cell free extract on commercial dyes was not successful, and only whole SRB cells resulted in decolourisation of the dyes. Consequently trials on the industrial dyes and effluents were carried out with whole cells. Decolourisation rates of up to 96 % were achieved for the commercial dyes and up to 93 % for the industrial dyes over a period of 10 days.
- Full Text:
- Authors: Mutambanengwe, Cecil Clifford Zvandada
- Date: 2007
- Subjects: Bioremediation , Dyes and dyeing -- Waste disposal , Sulfur bacteria , Hydragenase , Factory and trade waste -- Purification , Textile waste
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3960 , http://hdl.handle.net/10962/d1004019 , Bioremediation , Dyes and dyeing -- Waste disposal , Sulfur bacteria , Hydragenase , Factory and trade waste -- Purification , Textile waste
- Description: The continuing industrial development has led to a corresponding increase in the amount of waste water generation leading to a consequential decline in levels and quality of the natural water in the ecosystem. Textile industries consume over 7 x 10[superscript 5] tons of dyes annually and use up to 1 litre of water per kg of dye processed and are third largest polluters in the world, the problem being aggravated by the inefficiencies of the dye houses. An abundance of physio-chemical methods are in use world wide, however, there is increasing concern as to their impact in effectively treating textile effluents as they introduce secondary pollutants during the ‘remediation’ process which are quite costly to run, maintain and clean up. Research on biological treatment has offered simple and cost effective ways of bioremediating textile effluents. While aerobic treatment of textile dyes and their effluents has been reported, its major draw back is commercial up-scaling and as such anaerobic systems have been investigated and shown to degrade azo dyes, which form the bulk of the dyes used world wide. However, the mechanisms involved in the bioremediation of these dyes are poorly understood. The aims of this study were to identify and investigate the role of enzymes produced by sulphate reducing bacteria (SRB) in bioremediating textile dye and their effluents. Sulphate reducing bacteria were used in this study because they are tolerant to harsh environmental conditions and inhibit the proliferance of pathogenic micro-organisms. The appearance of clear zones in agar plates containing azo dye concentrations ranging from 10 – 100 mgl[superscript -1] showed the ability of SRB to decolourize dyes under anaerobic conditions. Assays of enzymes previously reported to decolourise azo dyes were not successful, but led to the identification of hydrogenase enzyme being produced by SRB. The enzyme was found to be localised in the membrane and cytoplasm. A surface response method was used to optimize the extraction of the enzyme from the bacterial cells resulting in approximately 3 fold increase in hydrogenase activity. Maximum hydrogenase activity was found to occur after six days in the absence of dyes but was found to occur after one day in the presence of azo dyes. A decline in hydrogenase activity thereafter, suggested inhibition of enzymatic activity by the putative aromatic amines produced after azo cleavage. Purification of the hydrogenase by freeze drying, poly ethylene glycol, and Sephacryl – 200 size exclusion- ion exchange chromatography revealed the enzyme to have a molecular weight of 38.5 kDa when analyzed by a 12 % SDS-PAGE. Characterisation of the enzyme revealed optimal activity at a pH of 7.5 and temperature of 40 °C while it exhibited a poor thermal stability with a half-life of 32 minutes. The kinetic parameters V[subscript max] and K[subscript m] were 21.18 U ml[superscript -1} and 4.57 mM respectively. Application of the cell free extract on commercial dyes was not successful, and only whole SRB cells resulted in decolourisation of the dyes. Consequently trials on the industrial dyes and effluents were carried out with whole cells. Decolourisation rates of up to 96 % were achieved for the commercial dyes and up to 93 % for the industrial dyes over a period of 10 days.
- Full Text:
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