Enzymatic recovery of rhodium(III) from aqueous solution and industrial effluent using sulphate reducing bacteria: role of a hydrogenase enzyme
- Authors: Ngwenya, Nonhlanhla
- Date: 2005
- Subjects: Enzymes , Rhodium , Enzymes -- Industrial applications , Sulfur bacteria , Hydrogenation , Hydragenase , Factory and trade waste -- Purification
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3956 , http://hdl.handle.net/10962/d1004015 , Enzymes , Rhodium , Enzymes -- Industrial applications , Sulfur bacteria , Hydrogenation , Hydragenase , Factory and trade waste -- Purification
- Description: In an attempt to overcome the high maintenance and costs associated with traditional physico-chemical methods, much work is being done on the application of enzymes for the recovery of valuable metals from solutions and industrial effluents. One of the most widely studied enzymatic metal recovery systems uses hydrogenase enzymes, particularly from sulphate reducing bacteria (SRB). While it is known that hydrogenases from SRB mediate the reductive precipitation of metals, the mechanism of enzymatic reduction, however, is not yet fully understood. The main aim of the present study was to investigate the role of a hydrogenase enzyme in the removal of rhodium from both aqueous solution and industrial effluent. A quantitative analysis of the rate of removal of rhodium(III) by a resting SRB consortium under different initial rhodium and biomass concentrations, pH, temperature, presence and absence of SRB cells and electron donor, was studied. Rhodium speciation was found to be the main factor controlling the rate of removal of rhodium from solution. SRB cells were found to have a higher affinity for anionic rhodium species, as compared to both cationic and neutral species, which become abundant when speciation equilibrium was reached. Consequently, a pH-dependant rate of rhodium removal from solution was observed. The maximum SRB uptake capacity for rhodium was found to be 66 mg rhodium per g of resting SRB biomass. Electron microscopy studies revealed a time-dependant localization and distribution of rhodium precipitates, initially intracellularly and then extracellularly, suggesting the involvement of an enzymatic reductive precipitation process. A hydrogenase enzyme capable of reducing rhodium(III) from solution was isolated and purified by PEG, DEAE-Sephacel anion exchanger and Sephadex G200 gel exclusion. A distinct protein band with a molecular weight of 62kDa was obtained when the hydrogenase containing fractions were subjected to a 10% SDS-PAGE. Characterization studies indicated that the purified hydrogenase had an optimum pH and temperature of 8 and 40°C, respectively. A maximum of 88% of the initial rhodium in solution was removed when the purified hydrogenase was incubated under hydrogen. Due to the low pH of the industrial effluent (1.31), the enzymatic reduction of rhodium by the purified hydrogenase was greatly retarded. It was apparent that industrial effluent pretreatment was necessary before the application an enzymatic treatment method. In the present study, however, it has been established that SRB are good candidates for the enzymatic recovery of rhodium from both solution and effluent.
- Full Text:
- Date Issued: 2005
- Authors: Ngwenya, Nonhlanhla
- Date: 2005
- Subjects: Enzymes , Rhodium , Enzymes -- Industrial applications , Sulfur bacteria , Hydrogenation , Hydragenase , Factory and trade waste -- Purification
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3956 , http://hdl.handle.net/10962/d1004015 , Enzymes , Rhodium , Enzymes -- Industrial applications , Sulfur bacteria , Hydrogenation , Hydragenase , Factory and trade waste -- Purification
- Description: In an attempt to overcome the high maintenance and costs associated with traditional physico-chemical methods, much work is being done on the application of enzymes for the recovery of valuable metals from solutions and industrial effluents. One of the most widely studied enzymatic metal recovery systems uses hydrogenase enzymes, particularly from sulphate reducing bacteria (SRB). While it is known that hydrogenases from SRB mediate the reductive precipitation of metals, the mechanism of enzymatic reduction, however, is not yet fully understood. The main aim of the present study was to investigate the role of a hydrogenase enzyme in the removal of rhodium from both aqueous solution and industrial effluent. A quantitative analysis of the rate of removal of rhodium(III) by a resting SRB consortium under different initial rhodium and biomass concentrations, pH, temperature, presence and absence of SRB cells and electron donor, was studied. Rhodium speciation was found to be the main factor controlling the rate of removal of rhodium from solution. SRB cells were found to have a higher affinity for anionic rhodium species, as compared to both cationic and neutral species, which become abundant when speciation equilibrium was reached. Consequently, a pH-dependant rate of rhodium removal from solution was observed. The maximum SRB uptake capacity for rhodium was found to be 66 mg rhodium per g of resting SRB biomass. Electron microscopy studies revealed a time-dependant localization and distribution of rhodium precipitates, initially intracellularly and then extracellularly, suggesting the involvement of an enzymatic reductive precipitation process. A hydrogenase enzyme capable of reducing rhodium(III) from solution was isolated and purified by PEG, DEAE-Sephacel anion exchanger and Sephadex G200 gel exclusion. A distinct protein band with a molecular weight of 62kDa was obtained when the hydrogenase containing fractions were subjected to a 10% SDS-PAGE. Characterization studies indicated that the purified hydrogenase had an optimum pH and temperature of 8 and 40°C, respectively. A maximum of 88% of the initial rhodium in solution was removed when the purified hydrogenase was incubated under hydrogen. Due to the low pH of the industrial effluent (1.31), the enzymatic reduction of rhodium by the purified hydrogenase was greatly retarded. It was apparent that industrial effluent pretreatment was necessary before the application an enzymatic treatment method. In the present study, however, it has been established that SRB are good candidates for the enzymatic recovery of rhodium from both solution and effluent.
- Full Text:
- Date Issued: 2005
The bioaccumulation of platinum (IV) from aqueous solution using sulphate reducing bacteria: role of a hydrogenase enzyme
- Authors: Rashamuse, Konanani Justice
- Date: 2003
- Subjects: Sulfur bacteria , Bioremediation , Enzymes -- Metabolism , Platinum , Platinum compounds , Reduction (Chemistry) , Hydrogenation
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4003 , http://hdl.handle.net/10962/d1004063 , Sulfur bacteria , Bioremediation , Enzymes -- Metabolism , Platinum , Platinum compounds , Reduction (Chemistry) , Hydrogenation
- Description: The enzymatic reduction of a high-valence form of metals to a low-valence reduced form has been proposed as a strategy to treat water contaminated with a range of metals and radionuclides. Metal reduction by sulphate reducing bacteria (SRB) is carried out either chemically (involving reduction by hydrogen sulphide) or enzymatically (involving redox enzymes such as the hydrogenases). While reduction of metal ions by hydrogen sulphide is well known, the enzymatic mechanism for metal reduction is poorly understood. The aims of this study were to investigate the role of SRB in facilitating platinum removal, and to investigate the role of a hydrogenase in platinum reduction in vitro. In order to avoid precipitation of platinum as platinum sulphide, a resting (non-growing) mixed SRB culture was used. The maximum initial concentration of platinum (IV), which SRB can effectively remove from solution was shown to be 50 mg.l⁻¹. Electron donor studies showed high platinum (IV) uptake in the presence of hydrogen, suggesting that platinum (IV) uptake from solution by SRB requires careful optimization with respect to the correct electron donor. Transmission electron microscopy (TEM) and energy dispersive X-ray (EDX) analysis indicated that platinum was being precipitated in the periplasm, a major area of hydrogenase activity in SRB. Purification of the hydrogenase by ammonium sulphate precipitation (65%), Toyopearl-Super Q 650S ion exchange and Sephacry 1 S-100 size exclusion chromatography revealed that the hydrogenase was monomeric with a molecular weight of 58 KDa, when analyzed by 12% SDS-PAGE. The purified hydrogenase showed optimal temperature and pH at 35°C and 7.5 respectively, and a poor thermal stability. In vitro investigation of platinum reduction by purified hydrogenase from mixed SRB culture showed that hydrogenase reduces platinum only in the presence of hydrogen. Major platinum (IV) reduction was observed when hydrogenase was incubated with cytochrome C₃ (physiological electron carrier in vivo) under hydrogen. The same observations were also noted with industrial effluent. Collectively these findings suggest that in vitro platinum reduction is mediated by hydrogenase with a concerted action of cytochrome C₃ required to shuttle the electron from hydrogenase.
- Full Text:
- Date Issued: 2003
- Authors: Rashamuse, Konanani Justice
- Date: 2003
- Subjects: Sulfur bacteria , Bioremediation , Enzymes -- Metabolism , Platinum , Platinum compounds , Reduction (Chemistry) , Hydrogenation
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4003 , http://hdl.handle.net/10962/d1004063 , Sulfur bacteria , Bioremediation , Enzymes -- Metabolism , Platinum , Platinum compounds , Reduction (Chemistry) , Hydrogenation
- Description: The enzymatic reduction of a high-valence form of metals to a low-valence reduced form has been proposed as a strategy to treat water contaminated with a range of metals and radionuclides. Metal reduction by sulphate reducing bacteria (SRB) is carried out either chemically (involving reduction by hydrogen sulphide) or enzymatically (involving redox enzymes such as the hydrogenases). While reduction of metal ions by hydrogen sulphide is well known, the enzymatic mechanism for metal reduction is poorly understood. The aims of this study were to investigate the role of SRB in facilitating platinum removal, and to investigate the role of a hydrogenase in platinum reduction in vitro. In order to avoid precipitation of platinum as platinum sulphide, a resting (non-growing) mixed SRB culture was used. The maximum initial concentration of platinum (IV), which SRB can effectively remove from solution was shown to be 50 mg.l⁻¹. Electron donor studies showed high platinum (IV) uptake in the presence of hydrogen, suggesting that platinum (IV) uptake from solution by SRB requires careful optimization with respect to the correct electron donor. Transmission electron microscopy (TEM) and energy dispersive X-ray (EDX) analysis indicated that platinum was being precipitated in the periplasm, a major area of hydrogenase activity in SRB. Purification of the hydrogenase by ammonium sulphate precipitation (65%), Toyopearl-Super Q 650S ion exchange and Sephacry 1 S-100 size exclusion chromatography revealed that the hydrogenase was monomeric with a molecular weight of 58 KDa, when analyzed by 12% SDS-PAGE. The purified hydrogenase showed optimal temperature and pH at 35°C and 7.5 respectively, and a poor thermal stability. In vitro investigation of platinum reduction by purified hydrogenase from mixed SRB culture showed that hydrogenase reduces platinum only in the presence of hydrogen. Major platinum (IV) reduction was observed when hydrogenase was incubated with cytochrome C₃ (physiological electron carrier in vivo) under hydrogen. The same observations were also noted with industrial effluent. Collectively these findings suggest that in vitro platinum reduction is mediated by hydrogenase with a concerted action of cytochrome C₃ required to shuttle the electron from hydrogenase.
- Full Text:
- Date Issued: 2003
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