Browsing by Author "Harrison, Susan T L"

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    Open Access
    A multifarious comparative ecotoxicological approach on a catchment scale from three mine wastelands for improved environmental management and risk assessment: integrating abiotic, biotic and agroecosystem approaches
    (2022) Mudenda, Lee; Harrison, Susan T L; Hudson-Edwards, Karen; Syampungani, Stephen
    Metal mobilization under acid or neutral rock drainage represents one of the major environmental impacts associated with mining of sulphidic minerals. To avert this, suitable handling of mine overburden material, waste rock, open pits and tailing storage facilities (TSF) is needed. This study addressed the risks associated with metal mobilization from the waste, including those resulting from the potential for ARD generation from the mine waste (tailings) following the recovery of copper from sulphidic ores, the impacts on the aquatic ecosystem, the agro-ecosystem and potential ecological restoration using phytomining technologies. The study is focused on the Kafue River basin on the Zambian Copperbelt and seeks to identify impacts and potential benefits through studying a grouping of TSFs and their impact in a single geographical region, allowing attributes of the facilities to be contrasted. In this study, we have addressed the categorisation of ARD generation of Chibuluma TSF, TSF15A and TSF14 tailing samples and associated metal mobility using the standard static tests, UCT biokinetic test and column bioleach experiments. Owing to the potential for compromised water quality and exploratory studies alluding to this, an ecotoxicology study at the catchment scale was conducted seasonally for three years on water resources (Nselaki Stream, Fikondo Stream and Mululu Streams) and food crops in close proximity to the selected TSFs. The potential of phytomining technologies using native herbaceous plants to mitigate mobilization of metals from the copper mine wastelands was investigated. Characterisation of the risk of ARD using data generated from the standard static and biokinetic tests was compared across the three samples. The biokinetic test supported the standard static test classification of non-acid forming, providing preliminary kinetic data on ARD generation. The three tailings have high neutralisation potential and are not acid forming over an initial period. Column bioleach tests allowed for differentiation of metals according to their leaching potential under conditions ranging from neutral through varying levels of acidity conditions, providing support evidence for potential ecological burdens. The results showed that low pH promoted significant release of Fe, Cu and Mn while release of metals Co, Ni, Zn and Pb remained considerably low. Low mobilization of metal species was observed under high pH, however, over time the sustained low mobilization of metal species is likely to cause significant ecological risks. The results better inform the risk posed by copper wastelands, through the combined use of a suit of tests (static, biokinetic and column leach tests). Under high acidic conditions, Fe and Cu exhibited high ecological risk while the risk was moderate under non-acidic conditions. The ecological risk under acidic conditions for Ca, Al, Mg and Mn was observed to vary from low to moderate, while negligible ecological risk profiles were observed with elements of interest Pb, Co, Zn, and Ni. Our research further expanded the studies on monitoring abiotic and biotic ecosystem drivers in adjacent streams. Selected physiochemical indicators downstream were identified in relation to the influence of the mine wastelands. No significant difference in heavy metals was observed between the three streams at the significance level (P > 0.05), however, notable changes in chemical and physical signatures for selected elements was reported downstream of the selected TSFs. Multivariate analysis such as principal component analysis, indicated prevalent TSF interferences of Cu, Co, Mn, Zn, and Pb in water and sediment samples analysed. The use of macroinvertebrates provided a useful approach to monitor the variation in the degree of impacts and characterise the ecological integrity of the streams, as well as evaluate the links with selected physiochemical contaminants. The various physiochemical markers used were useful in observing persistent impacts on macroinvertebrate taxa, which can be linked to severe anthropogenic impacts as well as timely warning indicators. Particularly, macroinvertebrate taxa tolerant to water pollution such as Talitridae and Gnathobdellidae were observed to be dominant species. The biotic monitoring results supported the abiotic test classification with regards to stream contamination. The use of macroinvertebrate community structures proved more useful to characterize the integrity of the ecosystem of the streams and determine the links with possible contaminants. Similarly, results from food crops irrigated using the selected streams reported significant elevation of metals Cu, Co, Mn, and Pb in the edible parts. The contamination load index (CLI) showed that the pollution index of Pb, at ≈43.8 in the vegetable samples, exceeded that of the other metals; equally, metal contamination was also determined in the edible vegetables for Cu, Co and Mn, but not consistently for Zn. One-way ANOVA at p≤0.05 and boxplot analysis suggests that heavy metal concentration in soils and crops did not vary significantly among the sites downstream of the TSF. In contrast, the soils in the upstream control sites showed much reduced metal content. These observations suggest that the TSFs may be the primary source of metal contamination in the selected streams. The study presents phytomining as an improvement approach towards mitigating the impacts of metal mobilization and rehabilitation of wastelands. Further, it acknowledges the benefit of vegetation of TSFs. A rich diversity of indigenous herbaceous plant species was observed to thrive on the low-grade wastelands, with 622 indigenous herbaceous species from 21 families and 46 genera identified. Through analysis of the rhizosphere and above- and belowground biomass of these plant species, the following plants reporting copper accumulation above 1000 ppm, terming them hyper-accumulators: A. eucomus, B. alata, C. floribunda, C. ductylon, C. alternifolius, H. filipendula, E. scuber and V. glabra. However, hyper-accumulation of Co, Zn and Mn was not observed despite accumulation to levels of 300, 200 and 1000 ppm respectively. Further, a number of the hyper-accumulators showed wide-spread acclimatisation to TSFs through their importance value index (IVI). Our findings suggest that phytomining using indigenous herbaceous plant species in Zambia has potential as a viable technology. Overall, the approach of comparing catchments impacted by similar land use activities, was observed to be valuable and useful in current and future management of watersheds exposed to similar challenges. The study highlights useful monitoring methods, key risks requiring mitigation and highlights the need for interventions. The comparative catchment scale study is unique and rare which few studies have utilised to assess the likely impacts of mine wastelands, while also investigating potential remedial measures.
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    Analysis of the microbial community associated with a bioprocess system for bioremediation of thiocyanate- and cyanide-laden mine water effluents
    (Trans Tech Publications, 2015-11) Huddy, Rob; Kantor, Rose; Van Zyl, Wynand; Van Hille, Robert P; Banfield, Jill; Harrison, Susan T L
    Gold extraction by cyanidation from refractory gold ores results in the formation of thiocyanate- and cyanide-contaminated wastewater effluents that must be treated before recycle or discard. Activated sludge processes, such as ASTERâ„¢, can be used for biodegradation of these effluent streams. The destruction of these compounds is catalyzed by a mixed microbial culture, however, very little is known about the community composition and metabolic potential of the thiocyanate- and cyanide-degrading microorganisms within the community. Here we describe our on-going attempts to better understand the key microorganisms, within the ASTERâ„¢ bioprocess, that contribute to the destruction of thiocyanate and cyanide, and how this knowledge relates to further process optimisation.
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    Attachment of Acidithiobacillus ferrooxidans and Leptospirillum ferriphilum cultured under varying conditions to pyrite, chalcopyrite, low-grade ore and quartz in a packed column reactor
    (Springer Verlag, 2012) Africa, Cindy-Jade; van Hille, Robert P; Harrison, Susan T L
    The attachment of Acidithiobacillus ferrooxidans and Leptospirillum ferriphilum spp. grown on ferrous medium or adapted to a pyrite mineral concentrate to four mineral substrata, namely, chalcopyrite and pyrite concentrates, a low-grade chalcopyrite ore (0.5 wt%) and quartzite, was investigated. The quartzite represented a typical gangue mineral and served as a control. The attachment studies were carried out in a novel particle-coated column reactor. The saturated reactor containing glass beads, which were coated with fine mineral concentrates, provided a quantifiable surface area of mineral concentrate and maintained good fluid flow. A. ferrooxidans and Leptospirillum spp. had similar attachment characteristics. Enhanced attachment efficiency occurred with bacteria grown on sulphide minerals relative to those grown on ferrous sulphate in an ore-free environment. Selective attachment to sulphide minerals relative to gangue materials occurred, with mineral adapted cultures attaching to the minerals more efficiently than ferrous grown cultures. Mineral-adapted cultures showed highest levels of attachment to pyrite (74% and 79% attachment for A. ferrooxidans and L. ferriphilum, respectively). This was followed by attachment of mineral-adapted cultures to chalcopyrite (63% and 58% for A. ferrooxidans and L. ferriphilum, respectively). A. ferrooxidans and L. ferriphilum exhibited lower levels of attachment to low-grade ore and quartz relative to the sulphide minerals.
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    Development of a method to assay the microbial population in heap bioleaching operations
    (Elsevier, 2005) Coram-Uliana, Nicolette J; van Hille, Robert P; Kohr, William J; Harrison, Susan T L
    Heap bioleaching is an economically viable approach to the mining of low-grade ores. Oxidation is microbially assisted, involving a consortium of microorganisms that together span the mesophilic to extreme thermophilic range of temperatures (25–80 °C). Temperatures inside the heap are not externally regulated, making the microbial interactions difficult to predict. In order to gain insight into these interactions, a qualitative and quantitative assay of the microorganisms that colonise the ore surface or are present in the liquid phase between the ore clusters at different levels within a heap has been developed. This method was developed using crude ore and liquid samples obtained from the GeoBiotics temperature controlled mesophilic heap operation at the Agnes Gold Mine in Barberton, South Africa, and the high temperature test columns at SGS Lakefield Research, Johannesburg, South Africa. This method of sample analysis may be applied to bioheap leach operations with and without temperature control. Ease of application, reproducibility and turn around time influenced technique design in order to provide a useful assay for commercial bioleaching operations. Following microbial removal from the solid phase using successive washes with detergent and acidified water, the cells were enumerated and genetic DNA was isolated. Microbial identification was achieved via restriction endonuclease analysis of the 16S rRNA genes, as well as 16S rRNA gene sequencing where necessary. Quantification was achieved using species-and genus-specific probes through fluorescent in situ hybridisation (FISH).
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    Effect of cell permeability and dehydrogenase expression on octane activation by CYP153A6-based whole cell Escherichia coli catalysts
    (BioMed Central, 2017-09-20) White, Bronwyn E; Fenner, Caryn J; Smit, Martha S; Harrison, Susan T L
    Background: The regeneration of cofactors and the supply of alkane substrate are key considerations for the biocatalytic activation of hydrocarbons by cytochrome P450s. This study focused on the biotransformation of n-octane to 1-octanol using resting Escherichia coli cells expressing the CYP153A6 operon, which includes the electron transport proteins ferredoxin and ferredoxin reductase. Glycerol dehydrogenase was co-expressed with the CYP153A6 operon to investigate the effects of boosting cofactor regeneration. In order to overcome the alkane supply bottleneck, various chemical and physical approaches to membrane permeabilisation were tested in strains with or without additional dehydrogenase expression. Results: Dehydrogenase co-expression in whole cells did not improve product formation and reduced the stability of the system at high cell densities. Chemical permeabilisation resulted in initial hydroxylation rates that were up to two times higher than the whole cell system, but severely impacted biocatalyst stability. Mechanical cell breakage led to improved enzyme stability, but additional dehydrogenase expression was necessary to improve product formation. The best-performing system (in terms of final titres) consisted of mechanically ruptured cells expressing additional dehydrogenase. This system had an initial activity of 1.67 ± 0.12 U/gDCW (32% improvement on whole cells) and attained a product concentration of 34.8 ± 1.6 mM after 24 h (22% improvement on whole cells). Furthermore, the system was able to maintain activity when biotransformation was extended to 72 h, resulting in a final product titre of 60.9 ± 1.1 mM. Conclusions: This study suggests that CYP153A6 in whole cells is limited by coupling efficiencies rather than cofactor supply. However, the most significant limitation in the current system is hydrocarbon transport, with substrate import being the main determinant of hydroxylation rates, and product export playing a key role in system stability.
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    Effect of culture conditions on the competition between lactate oxidisers and fermenters in a biological sulfate reduction system
    (Elsevier, 2012) Oyekola, Oluwaseun O; Harrison, Susan T L; van Hille, Robert P
    Kinetic constants (μmax and Ks) describing the predominance of lactate oxidation and fermentation were determined in chemostat cultures. The kinetics of sulfate reduction and lactate utilization were determined from 0.5 to 5 d residence times at feed sulfate concentrations of 1.0–10.0 g l−1. The kinetics of lactate fermentation in the absence of sulfate were investigated at residence times of 0.5–5 d. The lactate oxidizers (LO) were characterized by a μmax of 0.2 h−1 and Ks value of 0.6 g l−1 compared with a μmax of 0.3 h−1 and Ks of 3.3 g l−1 for the lactate fermenters (LF). Using mathematical models, it was shown that LO competed more effectively for lactate at low lactate concentrations (⩽5 g l−1) and high sulfide concentrations (0.5 g l−1). Lactate fermenters outcompeted the oxidizers under conditions of excess lactate (>5 g l−1) and low sulfide (0.014–0.088 g l−1).
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    Effect of culture conditions on the competitive interaction between lactate oxidizers and fermenters in a biological sulfate reduction system
    (Elsevier, 2012) Oyekola, Oluwaseun O; Harrison, Susan T L; Van Hille, Robert P
    Kinetic constants (lmax and Ks) describing the predominance of lactate oxidation and fermentation were determined in chemostat cultures. The kinetics of sulfate reduction and lactate utilization were determined from 0.5 to 5 d residence times at feed sulfate concentrations of 1.0–10.0 g l1 . The kinetics of lactate fermentation in the absence of sulfate were investigated at residence times of 0.5–5 d. The lactate oxidizers (LO) were characterized by a lmax of 0.2 h1 and Ks value of 0.6 g l1 compared with a lmax of 0.3 h1 and Ks of 3.3 g l1 for the lactate fermenters (LF). Using mathematical models, it was shown that LO competed more effectively for lactate at low lactate concentrations (65gl1 ) and high sulfide concentrations (0.5 g l1 ). Lactate fermenters outcompeted the oxidizers under conditions of excess lactate (>5 g l1 ) and low sulfide (0.014–0.088 g l1 ). 2011 E
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    Effect of inoculum size on the rates of whole ore colonisation of mesophilic, moderate thermophilic and thermophilic acidophiles
    (Elsevier, 2012) Tupikina, Olga V; Minnaar, Susanna H; Rautenbach, George F; Dew, David W; Harrison, Susan T L
    Bioheap leaching of low grade copper sulphides has been applied successfully at the commercial scale for the extraction of copper from secondary sulphide minerals. It is important to optimise the inoculation of heaps in order to minimise the residence time required for the heap and to maximise extraction.Thermophilic bioleaching of the primary sulphide chalcopyrite poses an additional challenge of rising temperatures inside the heap demanding microbial succession. After heap start up, rising heap core temperatures make conditions less favourable for mesophilic microbial species, and the moderately thermophilic community succeeds them in the consortium. In turn, thermophilic microorganisms succeed the moderately thermophilic microbes as the higher temperatures are reached.A detailed understanding of the microbial colonisation of whole ore is necessary to optimise microbial succession during thermophilic bioleaching, as is that of microbial growth kinetics on whole ore. Most published research is focused on microbial growth rates of bioleaching organisms in liquid cultures; little work is reported on microbial colonisation of whole ore and subsequent microbial activity. To extend the information available on the microbial diversity and succession in a bioleaching habitat, a study of bioleaching microbes colonising the ore body is required.The aim of this work was to explore aspects of colonisation of low grade chalcopyrite ore at 23 °C, 50 °C and 65 °C by acidophilic micro-organisms. Laboratory column packed bed reactors were designed to simulate heap leach environments and to provide a systematic way of studying microbial dynamics on whole ore. The effect of inoculum size and inoculation strategies on microbial activity established and the subsequent leaching performance were investigated under conditions that support mesophilic, moderately thermophilic and thermophilic microorganisms. A relationship was shown between the inoculum size and the culture time required to achieve Eh values greater than 700 mV, especially at 23 °C and 65 °C. However, the culture time required to establish an active iron- (and sulphur-) oxidising culture was also influenced by ore type, irrigation rate and inoculum adaptation. The effect on effluent Eh, pH and dissolved iron levels is also discussed.
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    The effect of nitrogen limitation on lipid productivity and cell composition in Chlorella vulgaris
    (Springer Verlag, 2014) Griffiths, Melinda J; van Hille, Robert P; Harrison, Susan T L
    Chlorella vulgaris accumulates lipid under nitrogen limitation, but at the expense of biomass productivity. Due to this tradeoff, improved lipid productivity may be compromised, despite higher lipid content. To determine the optimal degree of nitrogen limitation for lipid productivity, batch cultures of C. vulgaris were grown at different nitrate concentrations. The growth rate, lipid content, lipid productivity and biochemical and elemental composition of the cultures were monitored for 20 days. A starting nitrate concentration of 170 mg L−1 provided the optimal tradeoff between biomass and lipid production under the experimental conditions. Volumetric lipid yield (in milligram lipid per liter algal culture) was more than double that under nitrogen-replete conditions. Interpolation of the data indicated that the highest volumetric lipid concentration and lipid productivity would occur at nitrate concentrations of 305 and 241 mg L−1 , respectively. There was a strong correlation between the nitrogen content of the cells and the pigment, protein and lipid content, as well as biomass and lipid productivity. Knowledge of the relationships between cell nitrogen content, growth, and cell composition assists in the prediction of the nitrogen regime required for optimal productivity in batch or continuous culture. In addition to enhancing lipid productivity, nitrogen limitation improves the lipid profile for biodiesel production and reduces the requirement for nitrogen fertilizers, resulting in cost and energy savings and a reduction in the environmental burden of the process.
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    The effect of nitrogen limitation on lipid productivity and cell composition in Chlorella vulgaris
    (Springer Verlag, 2014) Griffiths, Melinda J; van Hille, Robert P; Harrison, Susan T L
    Chlorella vulgaris accumulates lipid under nitrogen limitation, but at the expense of biomass productivity. Due to this tradeoff, improved lipid productivity may be compromised, despite higher lipid content. To determine the optimal degree of nitrogen limitation for lipid productivity, batch cultures of C. vulgaris were grown at different nitrate concentrations. The growth rate, lipid content, lipid productivity and biochemical and elemental composition of the cultures were monitored for 20 days. A starting nitrate concentration of 170 mg L−1 provided the optimal tradeoff between biomass and lipid production under the experimental conditions. Volumetric lipid yield (in milligram lipid per liter algal culture) was more than double that under nitrogen-replete conditions. Interpolation of the data indicated that the highest volumetric lipid concentration and lipid productivity would occur at nitrate concentrations of 305 and 241 mg L−1 , respectively. There was a strong correlation between the nitrogen content of the cells and the pigment, protein and lipid content, as well as biomass and lipid productivity. Knowledge of the relationships between cell nitrogen content, growth, and cell composition assists in the prediction of the nitrogen regime required for optimal productivity in batch or continuous culture. In addition to enhancing lipid productivity, nitrogen limitation improves the lipid profile for biodiesel production and reduces the requirement for nitrogen fertilizers, resulting in cost and energy savings and a reduction in the environmental burden of the process.
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    The effect of nutrient supplementation on growth and leaching performance of bioleaching bacteria
    (Trans Tech Publications, 2009) van Hille, Robert P; Bromfield, L V; Botha, S S; Jones, G; van Zyl, A W; Harrison, Susan T L
    Heap bioleaching operations are often faced with extended and unpredictable lag periods after inoculation, prior to the establishment of a stable oxidising environment, during which the heap is fully colonised or the inoculum overcomes the sub-optimal conditions resulting from acid agglomeration. Supplementation of laboratory scale (4kg ore) leach columns with soluble nitrogen, particularly as yeast extract, significantly reduced the lag time and promoted bacterial growth, resulting in a 50-95% increase in copper recovery post-inoculation. The effect of yeast extract addition to Acidithiobacillus ferrooxidans in controlled oxidation tests was investigated. Initial exposure of a stock culture to yeast extract resulted in a transient, dose dependent inhibition at concentrations of 0.5 g.l-1 and below. At 1.25 g.l-1 inhibition was complete over the time scale of the experiment. The inhibition phase was characterised by observable changes in cell morphology and ultrastructure. Despite the initial inhibition, the biomass yield at the end of the experiments was equivalent, or higher, in the presence of yeast extract. Cultures were adapted to growth on yeast extract as the sole nitrogen source and adapted cultures showed the highest rates of iron oxidation and cell growth, in the presence of 0.5 and 1 g.l-1 of yeast extract.
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    Effect of sulphate concentration on the community structure and activity of sulphate reducing bacteria
    (Trans Tech Publications, 2007) Oyekola, Oluwaseun O; van Hille, Rob; Harrison, Susan T L
    This study investigated the effect of sulphate concentration and residence time on the performance of anaerobic sulphate reduction by a mixed sulphate reducing bacteria (SRB) culture using lactate as the sole carbon source and electron donor. The process perforrnance is related to the population structure of the microbial consortia and dominant metabolic reactions. Laboratory scale chemostat cultures at different residence times (1-4 d) and sulphate concentrations (1.0-10.0 glL) were employed. Lactate oxidation was prevalent at feed sulphate concentrations of 1.0 to 5.0 glL. A colresponding increase in the volumetric sulphate reduction rate with increasing volumetric loading rate was also observed at this range. However, at the higher feed sulphate concentration range (10.0-15.0 glL), sulphate inhibition, lactate fermentation and an increased microbial diversity were evident. At each feed concentration of sulphate in the range 5.0 to 15.0 glL, varying dilution rates resulted in significant shifts in dominant metabolic reactions. Sulphate concentration and residence time have significant effects on both the structure of the microbial population and kinetics of biological sulphate reduction.
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    Enhancing ARD mitigation by application of benign tailings to reduce the permeability of waste rock dumps
    (Trans Tech Publications, 2015-11) Kotsiopoulos, Thanos; Harrison, Susan T L
    ARD mitigation is considered by reducing the permeability of acid-generating coal interburden waste rock dumps. Fine desulfurised benign tailings were used to conceal exposed surface active sites from air, water and more aggressive sulfur- and iron-oxidizing microorganisms in acidic environments. Test columns containing interburden waste rock co-disposed with benign tailings were configured as either alternating layered covers or as a blended mixture of tailings and discards. Demonstration of the efficacy of the procedure was assessed by analyzing effluent solution concentrations of the concurrently operated simulated ore beds following inoculation with micro-organisms implicated in bioleaching (biotic mode) or in the absence of inoculation (abiotic mode). The presence of fine tailings both minimized the infiltration of oxidants to the porous ore and void space within the packed bed and provided neutralisation. In biotic mode, tests with layered covers of desulfurised tailings were less successful at limiting microbial activity and associated accelerated oxidation reactions. Blended columns resulted in reduced leaching reactions and an increase in the mean residence time of the solution phase. By increasing the solution residence time, an aqueous cover was concurrently formed, evolving into a bifunctional mitigating measure.
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    Evaluation of the ASTERTM process in the presence of suspended solids
    (Elsevier, 2014) van Zyl, Andries W; Huddy, Robert; Harrison, Susan T L; van Hille, Robert P
    The ability to recycle and reuse process water is a major contributing factor toward increased sustainability in the mining industry. However, the presence of toxic compounds has prevented this in most bioleaching operations. The ASTERTM process has been used for the bioremediation of cyanide (CN) and thiocyanate (SCN−) containing effluents at demonstration and commercial scale, increasing the potential for recycling of the treated effluent. The process relies on a complex consortium of microorganisms and laboratory tests have shown that the biomass retention, in suspended flocs or attached biofilm, significantly improved SCN− degradation rates. The current research evaluated the process performance in the presence of suspended solids (up to 5.5% m/v) ahead of implementation at a site where complete tailings removal is not possible. Experiments were performed in four 1 l CSTRs (with three primary reactors in parallel at an 8 h residence time, feeding one secondary reactor at a 2.7 h residence time). Stable operation at the design specifications (5.5% solids, 100 mg/l SCN− feed, effluent SCN− <1 mg/l) was achieved within 50 days, including a period of adaptation. The pH had the most significant effect on performance, with significant inhibition below pH 6. The presence of gypsum and anhydrite phases in the fresh tailings was most likely responsible for the observed decrease in pH. A maximum SCN− degradation rate of >57 mg/l/h was achieved, despite no obvious floc formation. Microbial ecology studies (16S rRNA clone library) revealed reduced diversity relative to reactors operated without suspended solids.
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    Feasibility for value addition to sucrose in South Africa through conversion to platform chemicals
    (2018) Jegede, Kemi; Harrison, Susan T L; Tai, Siew
    The world sugar price is constantly changing in response to supply and demand and is currently very low as compared to the prices it is sold at domestically in South Africa. The drop in the worldwide price of sugar is due to its oversupply as yields of sugar production have increased in recent years and subsidies and protection measures in other producing countries. The low prices also mean imports are cheaper than local sugar. This pushes down the average sugar price and leads to a low profit margin. Further, sugar production in South Africa is facing a number of challenges. The industrialization of the sugar belt in KwaZulu-Natal has resulted in less plantations and challenging topography for these. Incentivisation of small, medium and micro-scale commercial operations has increased the number of smaller scaled operations, with less economy of scale and less capital backing. Climatic factors have impacted crop yields. Production costs have increased in accordance with South Africa’s consumer price index whereas selling price has moved with the less inflationary global platform. Together, these have made the industry less economically viable. This has led to a need for value addition to sucrose and to eliminate the dependency on a single commodity. Re-positioning of sugar into value-added products has potential to boost the country’s economy by introducing other sources of revenue. Moreover there is a worldwide need to find alternative means to produce petroleum-based fuels and chemicals and bio-based products are being targeted to meet some of this need. A review of the global status shows that there has been value addition in the sugar industry producing mostly ethanol and other commodity chemicals such as surfactants, organic acids and polyols. It is therefore imperative to find sustainable ways of generating value added platform chemicals from sucrose. The quantitative and qualitative study of this project looks at determining the chemicals that should be considered as having the highest potential for value addition from sucrose in a South African context. The project was scoped to focus on chemicals and fuels that can be produced by biological conversions of sucrose. For the quantitative study, a set of 39 chemicals was selected from major studies performed globally on potential bio-based platform chemicals and these catalogued according to a set of criteria. The decision of the chemical/fuel to be studied was based on the gap in the chemical industry. This list comprised of chemicals that were selected in the US department of energy top 10 list in 2004 and 2010 and top 15 chemicals in the EU list in 2015. In addition to these, chemicals that are currently of interest (which were mostly chemicals that can be used as polymers and biofuels) were included to make up the list of 39 chemicals. The selected chemicals then went through a knock out selection where chemicals that cannot be produced with current technology from sugar or via a biological route were eliminated from the list. A quantitative analysis was then done on the remaining chemicals from the knock out stage. A weighting method which considered a series of factors was used to determine the top platform chemicals. The factors used were to identify platform chemicals that are at a high demand (both in South Africa and internationally), chemicals that showed great potential for profitability based on cost, technology readiness level and product yield. The quantitative analysis allowed seven chemicals to be selected. Finally a qualitative study based on interviews with experts in the field was done. Most of this information provided by the experts was supported by several literatures (Taylor, et al., 2015; Villadsen, et al., 2011; Choi, et al., 2015; Jansen & van Gulik, 2014). The qualitative study identified Succinic acid, Lactic acid and Citric acid as the top three chemicals. A techno-economic study was done on succinic acid, one of the most promising platform chemicals identified. The reasons for its selection was because it has a higher performance and it generates less carbon footprint than petroleum based succinic acid, competiveness for niche market, multiple application via BDO and PBS and its overall favourable environmental process that uses up carbon dioxide from the environment. Firstly, the succinic acid process was designed to be produced using Saccharomyces cerevisiae in a dual phase fed batch fermentation process. The overall design for the succinic acid process was based on the design proposed by Efe, et al (2013). A cost evaluation was then done on the design for an economic analysis. The economic analysis was done on the process to ascertain that there is indeed value addition of sucrose to the platform chemicals chosen. This was done in the form of profitability analysis of the process. An economic analysis of the design shows that the plant is profitable after the first year of operation. The total investment on the plant is R 22.3 billion and the start-up expense is R 1.05 billion. This project serves as a preliminary paper based overview of the general background for the selected platform chemicals that will be researched further in subsequent research.
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    The generation of toxic reactive oxygen species (ROS) from mechanically activated sulphide concentrates and its effect on thermophilic bioleaching.
    (Elsevier, 2011) Jones, Gavin C; Corin, Kirsten C; van Hille, Robert P; Harrison, Susan T L
    Two types of laboratory mills, planetary and vibratory, were used to activate sulphide mineral concentrates mechanically before thermophilic (bio)leaching. These samples were analysed in terms of particle size, surface area, density, SEM, XRD line profile analysis and reactivity. The product particle size distributions indicated different particle breakage mechanisms of the two mills. The surface area for pyrite milled with the planetary mill was three fold that milled in the vibratory mill for the same length of time. Planetary milled samples showed lower densities, up to 4% less for pyrite samples, compared to vibratory milled samples. Particle surface oxidation, observed by SEM, occurred post milling. Surface oxidation products were more prevalent with planetary milled sulphide samples. XRD line profile analysis showed more line broadening effects with the planetary mill. This indicated that more bulk particle-related structural defects were present in the planetary milled samples. The reactivity in acidic solution was measured in terms of the generation of toxic reactive oxygen species (ROS): hydrogen peroxide and hydroxyl radicals. The ROS generation from milled sulphides, normalised to constant surface area loading, increased with increased mechanical activation. The planetary milled samples generated greater ROS per sample surface area than vibratory milled samples, more than 4-fold for pyrite after 60 min of milling. Increased ROS generation was postulated to result from increased surface area defects, solubilisation of iron oxidation products and bulk particle-related defects. The effect of mechanical activation on performance on thermophilic leaching and bioleaching tests was investigated using milled samples at 2% (w/v) pulp density. Short mill times improved leach rates from both mills, up to 7-fold cf. unactivated feed leach rates. Poor bioleaching performance resulted following long periods of mechanical activation (20–60 min). Pyrite and chalcopyrite bioleaching performance decreased dramatically above surface area loadings of 25 and 125 m2 /L respectively. Planetary milled samples were less amenable to bioleaching. For pyrite milled for 20 and 60 min and chalcopyrite milled for 40 min, no viable cells were observed following inoculation via fluorescence microscopy, suggesting culture death supported by compromised ferrous iron oxidation. The generation of ROS was postulated to cause poor bioleaching performance under these conditions.
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    Improving the production of a thermostable amidase through optimising IPTG induction in a highly dense culture of recombinant Escherichia coli.
    (Elsevier, 2010) Olaofe, Oluwafemi A; Burton, Stephanie G; Cowan, Don A; Harrison, Susan T L
    The production of a novel thermostable amidase (EC 3.5.1.4) from Geobacillus pallidus RAPc8 using recombinant Escherichia coli BL21 (DE3) was investigated. Volumetric and specific enzyme activities were investigated in relation to inducer concentration in a batch process using a defined medium with glucose as the carbon source. While IPTG is routinely used to induce expression of genes under the control of lac promoter, the impact of high biomass concentration on IPTG induction has not been reported rigorously. In this study, biomass production was unaffected by IPTG concentration across the range 0–1000 M. Induction of recombinant protein expression by 400 M IPTG at late lag phase of growth (3rd hour) inhibited cell growth while induction at early exponential phase of growth (5th hour) gave a 3 fold increase in volumetric amidase activity compared to induction at mid exponential phase (8th hour). Protein production increased by a factor of two with IPTG addition, independent of IPTG concentration in the range of 40–1000 M. Amidase activity, measured on a volumetric basis and relative to protein and biomass concentrations, increased with increasing IPTG concentration up to 400 M. While inducer concentrations are typically reported on a volumetric basis, their mode of action is consistent with a biomass dependence. Analysis of the data across a range of biomass concentration confirmed that induction was a function of inducer concentration per unit biomass. The amidase enzyme was predominantly soluble and cytoplasmic with less than 3% retained within the cell debris.
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    In situ investigation and visualisation of microbial attachment and colonisation in a heap bioleach environment: the novel biofilm reactor
    (Elsevier, 2010) Africa, Cindy-Jade; Harrison, Susan T L; Becker, Megan; van Hille, Robert P
    In this paper, the development of a novel means of investigating the attachment and subsequent biofilm formation of mineral bioleaching micro-organisms to mineral surfaces in situ is described. The protocol was developed to investigate the interactions of micro-organisms with sulfide minerals and low-grade chalcopyrite ore under conditions resemblant of a bioheap environment. The method makes use of a biofilm reactor in which thin sections of mineral ore are mounted. The reactor is operated as a continuous flow-through system. Attachment of pure and mixed cultures of Acidithiobacillus ferrooxidans and Leptospirillum ferriphilum is assessed. The technique allows for the investigation of microbial ecology with special regard to microbe–mineral attachment, site and mineral specific associations of micro-organisms and spatial organisation of microbial communities present through the use of fluorescent microscopy techniques. Preliminary fluorescent in situ hybridisation (FISH) analysis of the attachment of L. ferriphilum and A. ferrooxidans to massive chalcopyrite sections, as well as to low-grade chalcopyrite containing ore sections is presented. In the case of both low-grade and massive sulfide mineral samples, attachment of mixed micro-colonies was observed in regions where surface defects were prevalent. In low-grade samples, preferential attachment was observed in regions where sulfide minerals were present. The density of the attached micro-colonies increased with an increase in contacting time (from 20, 72 and 96 h) and was indicative of an actively growing mono-layered biofilm.
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    The influence of microbial physiology on biocatalyst activity and efficiency in the terminal hydroxylation of n-octane using Escherichia coli expressing the alkane hydroxylase, CYP153A6
    (BioMed Central, 2013) Olaofe, Oluwafemi A; Fenner, Caryn J; Gudiminchi, Rama Krishna; Smit, Martha S; Harrison, Susan T L
    Biocatalyst improvement through molecular and recombinant means should be complemented with efficient process design to facilitate process feasibility and improve process economics. This study focused on understanding the bioprocess limitations to identify factors that impact the expression of the terminal hydroxylase CYP153A6 and also influence the biocatalytic transformation of n–octane to 1-octanol using resting whole cells of recombinant E. coli expressing the CYP153A6 operon which includes the ferredoxin (Fdx) and the ferredoxin reductase (FdR). Results: Specific hydroxylation activity decreased with increasing protein expression showing that the concentration of active biocatalyst is not the sole determinant of optimum process efficiency. Process physiological conditions including the medium composition, temperature, glucose metabolism and product toxicity were investigated. A fed-batch system with intermittent glucose feeding was necessary to ease overflow metabolism and improve process efficiency while the introduction of a product sink (BEHP) was required to alleviate octanol toxicity. Resting cells cultivated on complex LB and glucose-based defined medium with similar CYP level (0.20 μmol gDCW -1) showed different biocatalyst activity and efficiency in the hydroxylation of octane over a period of 120 h. This was influenced by differing glucose uptake rate which is directly coupled to cofactor regeneration and cell energy in whole cell biocatalysis. The maximum activity and biocatalyst efficiency achieved presents a significant improvement in the use of CYP153A6 for alkane activation. This biocatalyst system shows potential to improve productivity if substrate transfer limitation across the cell membrane and enzyme stability can be addressed especially at higher temperature. Conclusion: This study emphasises that the overall process efficiency is primarily dependent on the interaction between the whole cell biocatalyst and bioprocess conditions.
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    Interference by pigment in the estimation of microalgal biomass concentration by optical density
    (Elsevier, 2011) Griffiths, Melinda J; Garcin, Clive; van Hille, Robert P; Harrison, Susan T L
    Optical density is used as a convenient indirect measurement of biomass concentration in microbial cell suspensions. Absorbance of light by a suspension can be related directly to cell density using a suitable standard curve. However, inaccuracies can be introduced when the pigment content of the cells changes. Under the culture conditions used, pigment content of the microalga Chlorella vulgaris varied between 0.5 and 5.5% of dry weight with age and culture conditions. This led to significant errors in biomass quantification over the course of a growth cycle, due to the change in absorbance. Using a standard curve generated at a single time point in the growth cycle to calculate dry weight (dw) from optical density led to average relative errors across the growth cycle, relative to actual dw, of between 9 and 18% at 680 nm and 5 and 13% at 750 nm. When a standard curve generated under low pigment conditions was used to estimate biomass under normal pigment conditions, average relative errors in biomass estimation relative to actual dw across the growth cycle were 52% at 680 nm and 25% at 750 nm. Similar results were found with Scenedesmus, Spirulina and Nannochloropsis. Suggested strategies to minimise error include selection of a wavelength that minimises absorbance by the pigment, e.g. 750 nm where chlorophyll is the dominant pigment, and generation of a standard curve towards the middle, or across the entire, growth cycle.