Browsing by Author "Fagan-Endres, Marijke"
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- ItemOpen AccessDesulphurisation flotation for the selective removal of pyrite from coal discards using microorganisms(2018) Msipa, Winfull Jaconia; Harrison, Susan; Fagan-Endres, MarijkeMineral beneficiation processes such as base metal and coal mining produce large amounts of waste rock and coal discards that contain significant quantities of sulphide minerals with Acid Rock Drainage (ARD) generating potential. ARD is caused by the exposure of sulphide minerals, primarily pyrite (FeS2), to both water and oxygen, and microorganisms. This is a naturally occurring process, but the exposure of the sulphide containing mining wastes greatly accelerates ARD formation. Thus, ARD is a major issue associated with inactive mines, waste rock dumps and tailings impoundments, which over time presents a major environmental risk. The desulphurisation of coal discards, mine tailings and finely divided waste rock prior to their disposal has been proposed as a method of preventing ARD formation. This involves the selective separation of residual values from the waste rock, followed by selective separation of sulphide minerals – especially pyrite – from the residual waste material using a two-stage froth flotation to obtain a values stream, a low volume sulphide-rich concentrate that can be easily contained, and a high volume benign tailings fraction that can be safely disposed of. The technical feasibility of this two-stage process has been demonstrated; however, the cost of the flotation reagents used in this process are particularly high in comparison to the other operating costs, contributing as much as 75% of the operating costs for desulphurisation of coal fines. Furthermore, apart from being expensive, many of the inorganic flotation reagents are relatively toxic and could be hazardous to the environment due to their slow degradation rate. Microorganisms and their metabolic products have been identified in literature as potential reagents that can be used in the selective separation of sulphide minerals using froth flotation. Just like conventional chemical flotation reagents, the microorganisms assist separation through surface chemical alterations that modify a mineral’s hydrophobic properties, thus facilitating bioflotation. The aim of this study was to investigate the prevention of ARD formation through the desulphurisation of pyrite-containing coal discards and base metal hard rock samples using microbial cultures as alternative bioflotation reagents. In this study the feasibility of using P. polymyxa, R. palustris, R. opacus, B. subtilis, and B. licheniformis as biocollectors for the removal of pyritic sulphur in the second stage of the two-stage desulphurisation froth flotation process was investigated. Microbial screening tests were performed using a pyrite concentrate to assess each microbial culture’s affinity to pyrite and their ability to float the mineral in a batch flotation cell. Attachment experiments and batch bioflotation tests were carried out to screen for a microbial culture that showed potential. Following attachment experiments at pH 4 and pH 7, all microorganisms except B. licheniformis exhibited attachment to pyrite. The level of attachment was different for each microbial culture. P. polymyxa had the highest percentage attachment of 95.6 ± 1.0 % at pH 4 and 97.1 ± 0.7 % at pH 7 after 20 minutes of interaction. Subsequent results from the pyrite-only bioflotation tests revealed that R. opacus, R. palustris and B. subtilis did not affect the floatability of pyrite. P. polymyxa, however, showed a significant effect on the floatability of pyrite, achieving a cumulative mass recovery of 7.0 ± 0.42 % at pH 4 and 81.3 ± 0.4 % at pH 7. Zeta-potential tests revealed that P. polymyxa had the most neutral net surface charge across the pH range tested, while the other microorganisms had a large net positive or negative charge. Based on this result, it was deduced that the hydrophobicity of P. polymyxa as a consequence of its near neutral surface strongly made it seek out a surface to attach to rather than remaining suspended in water. Hence, P. polymyxa was chosen as the bio-collector candidate for the bioflotation separation of pyritic sulphur from coal discard and base metal hard rock samples. Despite the positive batch pyrite bioflotation tests, P. polymyxa was not successful for the flotation of pyrite from the coal discards nor did it upgrade pyritic sulphur to the concentrate, with the bioflotation results not significantly different from the negative control without collector. P. polymyxa did affect the floatability of the base metal hard rock, achieving cumulative mass recoveries comparable with the chemical control using PAX. However, there was no significant upgrade of pyritic sulphur content, with the biofloat achieving 22.6 % total sulphur in the concentrate which was significantly less than the 66.4 % total sulphur recovered with PAX. The study thus yielded positive results from fundamental studies of P. polymyxa’s ability to enhance the flotability of pyrite. However, tests using actual samples were less successful. Although P. polymyxa enhanced the floatability of the base metal hard rock, it did not achieve the aim of obtaining a low volume sulphide-rich concentrate as the PAX did. Recommendations for the continuation of this work include contact angle measurements and FT-IR spectroscopy to better understand the effects of P. polymyxa attachment, as well as performing a kinetic study on the growth of P. polymyxa alongside adaptation of the microbial culture to a pyrite mineral concentrate in order to test if this can improve selective flotation of the desired mineral owing to modified surface properties.
- ItemOpen AccessDesulphurisation of fine coal waste tailings using algal lipids(2018) Chiodza, Kudzai Godknows; Harrison, Sue; Fagan-Endres, MarijkeThe South African economy is an energy-driven economy which relies on coal to meet most of its energy demands. Coal mining has resulted in the generation of coal waste over 60 million tonnes, annually. Apart from the huge footprint of this waste, the sulphide minerals contained in the waste have resulted in the generation of acid rock drainage (ARD). A lot of techniques have been developed to prevent and mitigate ARD, however most of these techniques have fallen short in terms of meeting their desired objectives due to the long-term nature of ARD generation which can persist for hundreds of years after mine closure. This has resulted in emphasis being put on long-term prevention techniques that remove ARD risk over treatment techniques. One prevention technique which has shown good technical potential is the two-stage flotation method developed for desulphurisation of hard rock tailings and coal fines, developed at the University of Cape Town. On desulphurising coal, the first stage produces an upgraded coal product that may be sold, with the second stage used to separate the tailings from the first stage into targeted high-sulphide and low-sulphide fractions which may then be appropriately used or disposed of. An economic assessment of the process showed across a wide range of coal wastes the high cost of oleic acid used in the first stage of the process as a collector was a major contributor to the operating costs. The investigation undertaken in this thesis looked at the potential of algal lipids and their derivatives as biocollectors to replace the oleic acid collector in the desulphurisation process at the laboratory scale. A review of cost was carried out for a process that used raw algal lipids (RALs) or fatty acid methyl esters (FAMEs), which are derived from RALs through transesterification. Batch flotation experiments were used to assess the performance of the two bioflotation reagents in comparison to oleic acid and dodecane, an alternative but less successful chemical collector. The algal lipids cost review was a desktop study which was done by adapting literature data from Davis et al. (2014) which focused on economic evaluation of algal lipid biofuels production pathways. Results from laboratory experiments for two different coal waste feed samples showed that the performance of RALs and FAMEs was similar to that of oleic acid for the sample that was high in ash and sulphur, and better than oleic acid for the sample that was low in ash and sulphur. For example, the product from Site 1 discards from Waterberg had 24.37% ash and 2.76% sulphur using FAMEs, 26.13% ash and 2.56% sulphur with RALs, and 23.48% ash and 2.41% using oleic acid, at a reagent dose of 2.8 kg/t for all reagents. For Site 2 waste tailings from the Witbank area, the product had 23.17% ash and 0.72% sulphur when FAMEs were used as collector, 22.75% ash and 0.75% sulphur with RALs, and 20.18% ash and 0.74% sulphur using oleic acid, at the same reagent dose. Discards from Site 1 had an initial ash and sulphur content of 47.61% and 5.71%, respectively. Site 2 waste tailings had 25.56% ash and 0.91% sulphur before flotation. Increasing biocollector dosage resulted in higher yields with a compromise on the upgraded coal quality. The pH tests showed that the performance of the two bioflotation reagents was best at pH 4 in terms of yield. However, increasing the pH of the process from the natural pH of the sample (pH 2.7) to 7 resulted in collection of more ash and sulphur, thus reducing the product quality. The algal lipids cost review showed that RALs and FAMEs were potentially 20 to 21% cheaper than oleic acid, with more room for improvement. Both the laboratory experiments and the technical evaluation showed that algal lipids and their derivatives have the potential to replace oleic acid in the two-stage desulphurisation process for coal waste to obtain a saleable quality coal product while simultaneously decreasing the impact of ARD from coal waste.
- ItemOpen AccessDeveloping quantitative approaches to determine microbial colonisation and activity in mineral bioleaching and characterisation of acid rock drainage(2019) Makaula, Didi Xhanti; Harrison, Susan; Fagan-Endres, Marijke; Huddy, RobertColonisation of mineral surfaces by acidophilic microorganisms during bioleaching is important for accelerating the extraction of valuable metals from mineral sulfide ores of varying grades through biohydrometallurgy. It also influences acid formation and mineral deportment from sulfidic waste rock generated in mining processes and is key to its comprehensive waste rock characterisation for acid forming potential. This study assesses mixed mesophilic microbial interactions with, and colonisation of, pyrite concentrates and pyrite bearing waste rocks. The assessment of these interactions was carried out in this study in a synergistic qualitative as well as quantitative manner, with a particular focus on heap bioleaching for metal extraction and on disposal of waste rock, the latter through the case of characterisation of ARD generation potential. Using the tools developed, both the course of colonisation and development of metabolic activity with time of colonisation, as well as their correlation with leaching performance were studied. Furthermore, specific operating parameters such as ore grade and irrigation rates were explored. Finally, the application of this knowledge in a characterisation study was explored. To achieve the set of tools required for this study, two quantitative techniques were refined to characterise these microbial-mineral interactions. In the first, an isothermal microcalorimetric (IMC) method was developed and optimised to determine microbial colonisation of mineral surfaces quantitatively as a function of surface area (m-2 ). Three IMC configurations were considered: colonised pyrite-coated beads submerged in fresh media; beads submerged in cell free leachate; and beads in an unsaturated bed, each in the IMC vial. The highest heat output was measured in the unsaturated bed (263.3 mW m-2 ). The consistency of heat produced by the colonising microorganisms was determined through reproducibility studies. Using IMC, chemically and microbially facilitated pyrite oxidation rate studies were performed on unsaturated beds with varying surface area loadings, correlating to varying bead number. Results obtained showed similar normalised oxidation rates per surface area across the surface loadings. However, with more microbially colonised surface area loaded, the maximum heat generated was reached more quickly. This suggested that there was reagent (possibly O2) limitation in the system, which restricted microbial activity and its associated heat generation. Reagent limitation in the system was tested and validated through varying the O2 availability in the IMC vial by air displacement with CO2 and N2 gas, with the systems containing less O2 showing limited activity. Collectively the data showed that high activity, facilitated microbially, was achieved in unsaturated systems in a reproducible manner. Secondly, oxidation rates were determined and O2 limitation in the system was overcome. This then fundamentally informed the determination of activity from microbial-mineral interaction, using IMC, as a function of surface area. Secondly, a detachment protocol developed at UCT to recover microbial cells from surfaces of crushed and agglomerated ore to assess microbial growth rates and distribution in the ore bed, including cells in the interstitial phase and those weakly and strongly attached to the ore surface, was refined to assess colonisation of the finely milled pyrite-bearing concentrate or waste rock coated onto glass beads in continuous flow assays. The detachment protocol was assessed quantitatively by measuring initial and residual microbial activity, as a function of wash number, using IMC, thus providing a new level of confidence in the method. Mineral surfaces were visualised using scanning electron microscopy (SEM) following detachment for qualitative assessment. These data, together with microscopic enumeration of detached cells with increased number of washes, allow refinement of the assay and showed that six washes provided reliable estimation of mineral associated microbial cells. Extracellular polymeric substances (EPS) produced in this process were extracted using crown ether and the capsular bound components analysed. The analysed components included lipids (4.2 %), iron (16.4 %), DNA (26.8 %), and total carbohydrates (28.5 %), which are typical components of EPS. The carbohydrate fraction was further resolved to trehalose (26.2 %), fructose (36.5 %) and galactose (37.3 %) sugar monomers. The analysed EPS components confirmed presence of the EPS secreted by cells colonising the mineral ore or waste rock surface in a flow-through system, and visualised via SEM. The microcalorimetric approach developed together with the refined detachment method were applied to samples from a flow-through mini-column system, used to simulate microbe-mineral contacting in a heap. Here, the colonisation of pyrite concentrate by a mixed mesophilic culture of iron and sulfur oxidising microorganisms was assessed progressively over 30 days. The progression of mineral colonisation in the mini-column system was monitored using a combination of IMC, scanning electron microscopy, detachment method and conventional wet chemistry measurements. We observed an increase in the heat output from the colonised surfaces of pyrite mineral concentrate caused by oxidative reactions facilitated by mineral-microbial biofilm. This confirmed that the attached microorganisms were metabolically active and facilitated ongoing mineral leaching through regeneration of lixiviants. Correlation was shown between number of cells detached from the mineral surface and the heat generated, with a constant heat output per cell observed until day 15 of operation. Thereafter, the measured heat generated per cell increased, suggesting reduced efficiency of cell detachment owing to increasing firm attachment, or the lack in separation of single cells embedded within EPS matrix (clumps observed under light microscope after detachment). Using IMC to quantify the activity of the residual microorganisms on the mineral surface following detachment, it was confirmed that >95% of activity was detached through this protocol, hence the lower detached cell numbers determined following EPS formation were attributed to clumping of the detached cells. This correlated to an increased presence of EPS and was supported by SEM observation. Following the study of pyrite concentrate, colonisation of two pyrite bearing waste rock samples was assessed, with simultaneous establishment of the flow-through mini column biokinetic test configuration that resembles open flow in the waste rock dump. The flowthrough configuration was run alongside the refined UCT-developed batch biokinetic test using suspended mineral. In this study, two pyritic waste rock samples, liberated by milling, were characterised using three biokinetic test approaches: the slurry batch test (BT), the batch test using mineral-coated beads (BT-CB) and flow-through column test with mineral-coated beads (FT-CB). Our results have shown through static tests, solution redox potential and pH analysis that both waste rocks were acid forming. Furthermore, it was demonstrated in the FT-CB system that microbial proliferation on the waste rock surfaces progressed with time such that oxidative exothermic reactions facilitated by the increasing microbial presence on the surfaces were demonstrated using Isothermal microcalorimetry. This study presents and informs the on-going refinement of the biokinetic test through establishment of a flow-through test for ARD characterisation while providing insight into the role of the microbial phase in ARD generation. Microbial-mineral association was assessed under various operating conditions, including two solution flow rates (60 and 4 ml h -1 ) and minerals of varying sulfide content, including a pyrite concentrate (96 % pyrite), a high sulfide waste rock (33 % pyrite) and a low sulfide waste rock (14 % pyrite). Mineral grade impacted the activity of mineral associated microorganisms with higher activities observed on a mineral surface with high sulfide content. The activity measured from microorganisms that were associated with the pyrite concentrate was 827 mW m-2 at a 60 ml h -1 flow rate, whereas activity measured on low and high sulfide waste rock (PEL-LS and PEL-HS) were 293 mW m-2 and 157 mW m-2 respectively operated on the same flow rate. On decreasing the flow rate to 4 ml h -1 , the activity of microbial cells on PEL-LS and PEL-HS were 153 mW m-2 and 146 mW m-2 respectively. This study showed that the growth of microbial cell numbers coupled with metabolic activity is important to facilitate accelerated dissolution of sulfidic mineral surfaces. The rate of oxidation increased in the presence of EPS and thus EPS was further analysed, and its composition was confirmed. Overall, this study contributed to the understanding of microbial colonisation of mineral surfaces in a non-destructive quantitative manner. This study thus demonstrates the ability to measure and track both the growth and activity of microorganisms that are associated with mineral surfaces. This is important as it provides an approach to understanding microbe mineral surface interactions and, therefore, potential strategies to increase microbial colonisation of low-grade minerals that house valuable metals, during commercial heap bioleach processes. Furthermore, the ability to monitor progressive growth and activity of mineral associated microbial communities within a flow-through biokinetic test, as successfully demonstrated in this study, has the potential to significantly enhance current management of mine waste materials and ARD mitigation strategies. Therefore, on-going investigations of progressive microbe-mineral interactions will continue to be valuable both in terms of bioleaching for metal recovery and the mitigation of ARD through effective characterisation of mine waste material.
- ItemOpen AccessFermented foods production using isolated Lactobacillaceae species for the improvement of vaginal health: the case of mageu and yoghurt(2022) Hartzenberg-Aeroe, Ellen; Fagan-Endres, Marijke; Kullin, BrianBacterial vaginosis (BV) is the most prevalent vaginal dysbiosis affecting women's reproductive health. The condition is characterised by the disruption of a Lactobacillus spp.-dominated microbiota that is associated with positive health outcomes and a shift to a microbial community that consists of diverse obligately anaerobic bacteria. Currently, antibiotics are used to treat BV and other vaginal infections. However, post-therapy recurrence and reinfection rates are high, suggesting the need for adjunctive therapy. Probiotic supplements can be used in conjunction with antibiotic treatment to re-establish the optimal vaginal microbial community. Of the probiotic products available in South Africa, only a limited number are specifically for vaginal health and none contain probiotic microorganisms found in the female genital tract. Additionally, the healthy vaginal microbiota differ with geography. Therefore, there is a need for an affordable probiotic product targeted for South African women. In this dissertation, mageu and yoghurt are investigated for their potential as probiotic delivery vehicles. Previously isolated Lactobacillaceae strains from the genital tracts of healthy South African women are tested for their ability to ferment maize meal to mageu and milk to yoghurt, both as pure cultures and supplemented with traditionally-used microorganisms: Saccharomyces cerevisiae for mageu and Streptococcus thermophilus for yoghurt. Protocols were developed for the production of mageu and yoghurt with the bacterial strains. During production, fermentation was monitored by measuring pH until the end points of 3.5 and 4.5 were reached for mageu and yoghurt, respectively. After fermentation, the products were analysed by measuring titratable acidity, lactic acid and ethanol concentrations, total solids content, viable cell counts, qualitative analysis and shelf-life. Four mageu samples were then analysed for their consumer acceptability by an untrained consumer panel. The Lactobacillaceae isolates, both as pure strains and supplemented with S. cerevisiae, were able to ferment maize meal to mageu, producing a product with a final pH of around 3.5, which qualitatively resembled mageu. The presence of pH-lowering metabolites including lactic acid and other acids was also measured using titratable acidity. The addition of S. cerevisiae significantly reduced the fermentation time and increased the titratable acidity of all mixed culture samples compared to the pure culture mageu samples. All mageu samples produced using the Lactobacillaceae isolates met the ethanol (maximum 2.5 g/L) and total solids content (minimum 8% (m/m)) production requirements set by the South African National Standards. After one week at room temperature or refrigerated, none of the mageu samples had pH values below 3, which is acceptable for mageu. However, the mixed culture mageu samples saw a larger drop in pH over the week than the pure culture samples. The sensory analysis investigated the consumer acceptability of four probiotic mageu samples, a positive control produced with flour as the inoculum source, and a commercial sample. The commercial sample was generally preferred, and was scored highest by the consumer panel, while the other samples received mixed results. The spread of results was attributed to the influence of individual preference due to the small cohort size. The pure Lactobacillaceae strains were unable to ferment milk to yoghurt. By hour 12, the pH values were still between 6.32 and 6.37, and curdling of the product had occurred by hour 24. However, when supplemented with S. thermophilus, the mixed cultures were able to produce a product with a final pH of around 4.5 that qualitatively resembled traditional yoghurt. However, it is not known if the improved fermentation was due to S. thermophilus activity alone. The titratable acidity results also confirmed that pH-lowering metabolites were produced and all yoghurt samples had titratable acidity values greater than 0.6% (m/m), the minimum requirement for yoghurt. Little difference was observed between the samples for the qualitative assessment; however, the positive control had a thicker consistency than the probiotic samples. These results show that the Lactobacillaceae isolates when supplemented with traditional fermentation microorganisms are able to produce mageu and yoghurt. During this project protocols for mageu and yoghurt production using the Lactobacillaceae isolates specific to South African women were designed. This project serves as the first step towards investigating the use of fermented foods as affordable probiotic delivery vehicles for the improvement of vaginal health in South African women.
- ItemOpen AccessImpact of cryoprotectants during freeze drying on Lactobacillus plantarum viability and their role in enhancing probiotic storage stability(2021) Oluwatosin, Olasumbo O; Fagan-Endres, Marijke; Tai, SiewThe human microbiome has recently garnered the interest of scientists and biopharma industries as studies have revealed the potential use of live bacteria known as probiotics as potential therapeutics for restoring and maintaining human health. These probiotic-biopharma formulations must contain the right strain(s) in sufficient numbers when administered to confer the desired health benefit. Cell dehydration is used to keep the probiotic microbes in an inactivated form during storage, thereby ensuring that there are enough viable cells still present when the probiotic is taken. However, the drying process itself is detrimental to the probiotic cells and can result in reduced viability and stability of cells over storage. In this study, various cryoprotectants were assessed for their ability to maintain cell integrity and improve yield during the freeze drying dehydration of Lactobacillus plantarum towards a potential topical pharmabiotic formulation. Inulin, sucrose, maltodextrin, and skimmed milk at 10% m/v concentration of the drying media were tested for their ability to protect bacterial cells during freeze drying and over a storage period of 12 weeks at 4oC and room temperature. Furthermore glucose, inulin, sucrose, and maltodextrin as sole carbon substrate were investigated as prebiotics in concentrations of 0.5% m/v, 2% m/v, and 4% m/v of the fermentation media by in vitro fermentation of L. plantarum in glucose-free MRS-free media. The influence of these cryoprotectants and prebiotics on L. plantarum was measured against cell viability, growth kinetic parameters (growth rate, lag phase, and maximum cell density), and pH reduction potential of L. plantarum. Improved survival of L. plantarum during freeze drying and over 12-weeks of storage was observed with all cryoprotectants. Skimmed milk demonstrated the highest protection after freeze drying, with a survival rate of 91% and viable cell counts of 9.1 × 108 ( CFU ml ) from an initial cell count prior to drying of 1.0 × 109 ( CFU ml ). Inulin demonstrated high protective efficiency, with 85% viability maintained during freeze drying which resulted in final cell counts of 1.1 × 109 ( CFU ml ) from an initial cell count of 1.3 × 109 ( CFU ml ). However, inulin provided the least protection over the 12 week storage period compared to cells dried in the presence of maltodextrin, sucrose, and skimmed milk, with cell counts of only 1.2 × 106 ( CFU ml ) at 4oC and 6.3 × 103 ( CFU ml ) at room temperature recorded at the end of the period. Following skimmed milk, which also demonstrated the highest stability of cells over storage, sucrose performed second best in maintaining the stability of cells at 4 oC at the end of the 12 weeks storage, with viability of 33% which resulted in final cell counts of 3.4 × 108 ( CFU ml ). Overall, the presence of cryoprotectants and prebiotics demonstrated a significant influence on propagation and viability. The presence of each of the various prebiotics as the sole carbon substrate in the fermentation media promoted proliferation of L. plantarum. An increase in cryoprotectant concentrations led to increased biomass yield but with no significant change in the growth rate and lag phase. Cells showed improved stability when stored at 4oC compared to room temperature. A delay in propagation up to 10 hours was observed upon rehydration of stored probiotic cells across all cases except for skimmed milk that resulted in a maximum delay in propagation of 2 hours at both storage temperatures.
- ItemOpen AccessImproving an aqueous two-phase process for C-phycocyanin extraction from Spirulina(2022) Hockey, James Temlett; Harrison, Susan; Fagan-Endres, MarijkeBiotechnology and bioprocess engineering have made it possible to expand production of natural compounds, and markets have moved more towards this direction. An example of this is in pigments, where many synthetic pigments have been banned or pulled out of the market due to health concerns, while naturally-derived pigments are growing in popularity. The algal pigment, C-phycocyanin (C-PC) is a blue photosynthetic pigment of the phycobiliprotein family, found in cyanobacteria and red algae. Algal pigments like C-PC have been growing in demand and the market is expected to grow at 5-7 % annually. Phycocyanin has applications as a food and cosmetics dye, a health product with therapeutic uses, and as a diagnostic protein. Various processes have been studied to recover and purify C-PC from cyanobacteria such as Arthrospira platensis, commonly referred to as Spirulina, the most used organism for producing the pigment. The C-PC recovery process includes extraction, recovery and purification steps. One recovery and purification step reported in literature is aqueous twophase separation (ATPS), which is able to produce high purity C-PC with good recovery. At the University of Cape Town (UCT), the Centre for Bioprocess Engineering Research (CeBER) has patented a process for extracting and purifying C-PC from Spirulina using a polyethylene-glycol (PEG) and maltodextrin (MDX) ATPS. This is a less-studied form of ATPS, with most C-PC extraction studies using PEG – salt systems. The PEG – MDX system was studied due to challenges faced with C-PC recovery from the PEG phase. The patented CeBER process begins with cell disruption, a period of leaching into a buffered solution followed by cell debris removal. The C-PC is subsequently purified and recovered by the PEG – MDX ATPS and three ammonium sulfate precipitation stages, and finally dried to powder. The patented process requires optimisation and up-scaling before being commercially applied in industry, the ultimate aim of the greater project. As such, this project aimed to improve the understanding of the overall process for C-PC recovery from Spirulina and, in particular, the ATPS step involved, to improve the ATPS performance. The study also sought to produce cosmetic grade C-PC (purity number of > 1.5), and develop process options and simulations for this production based on a combination of literature and experimental results. The work was conducted in view of future up-scaling. This included study of the leaching step with the Spirulina used in this project as well as refinement and optimisation of the ATPS step. For the latter, phase diagrams for the PEG – MDX ATPS were produced to inform ATPS refinement and improvement before evaluating its performance. The phase diagrams give an understanding of how the phases partition in an ATPS, allowing prediction and optimisation of top and bottom phase compositions. Leaching experiments showed that a maximum C-PC concentration was found after 2 h with the Spirulina powder used without the need for cell disruption, and that the purity decreased slowly over time (using 100 g/L Spirulina powder in 5 g/L citrate buffer at pH 6). This recommended a relatively short leaching time be used compared to previous work using different starting material. The PEG – MDX ATPS phase diagrams produced corresponded well with similar ATPS data found in literature. The PEG molecular weight was tested for the best performance, finding that PEG 10000 performed slightly better than both PEG 6000 and PEG 20000, achieving a C-PC purification factor of 1.21 ± 0.01 at 9 wt% PEG 10000 and 20 wt% MDX, with a recovery of 95.1 ± 7.8 %. However, the PEG – MDX ATPS for C-PC purification gave lower purification factors compared to PEG – salt ATPS studies from literature. A two-stage ATPS was therefore considered, with a PEG – citrate ATPS used before the PEG – MDX ATPS. This aimed to take advantage of the good C-PC selective recovery reported in literature for PEG – salt ATPS systems while still using the PEG – MDX to separate the C-PC from the PEG phase. PEG – citrate phase diagrams were produced; these compared well with those found in literature studies. A screening of PEG molecular weights across both ATPS steps found PEG 4000 to be best, mainly due to the performance in the PEG – citrate stage. Using PEG 4000 in two factorial studies on the impact of PEG and citrate concentrations in the first ATPS, and PEG and MDX concentrations in the second ATPS, response curves for the purification, recovery and concentration of the C-PC in the desired phase were produced. A Statistica (version 13.5.0.17) model was used to predict a local optimum, where the combination of component concentrations produce C-PC at high purify, recovery and concentration. The PEG – citrate ATPS model predicted the best component concentrations to be 11 wt% PEG and 20 wt% citrate, which gave a C-PC purification factor of 1.63 ± 0.28, at a recovery of 95.6 ± 8.0 %. The PEG – MDX ATPS model predicted a 1.43 ± 0.09 C-PC purification factor and a recovery of 86.8 ± 4.7 %, using a composition of 11 wt% PEG and 22 wt% MDX. A combination of experimental results and literature data were used to underpin the simulation of five process configurations for C-PC production using SuperPro Designer (version 9.5), each targeting a cosmetic grade C-PC product or better. These sought to simplify and improve C-PC production using the PEG – MDX ATPS as the core unit procedure. The first simulation was of the original patented process (cell-disruption, leaching, ATPS, precipitation and finally freeze-drying), with the ATPS operation updated with the best case experimental results obtained in this work. The second and third simulations used the newly proposed two-stage ATPS. In the third option ultrafiltration replaced the precipitation steps. Spray-drying replaced freeze-drying as a faster and more cost-effective means of drying C-PC from the second simulation onward. The fourth simulation incorporated a pre-treatment step, using activated carbon and chitosan to adsorb contaminant proteins and purify the C-PC, before using a single PEG – MDX ATPS. The operation and performance of the pre-treatment step were based on literature information. This model used (NH4)2SO4 precipitation as in the original process and required two precipitation stages for final purification. The fifth simulation used the pre-treatment process as in simulation 4, with the second precipitation step replaced with filter-sterilisation, before spray-drying. The two-stage ATPS processes lead to slightly improved recoveries of 39.1 % and 39.5 % for the second and third of process recommendations, respectively. This is above the original process simulated to recover 38.7 % of the C-PC in the crude starting solution. The two-stage ATPS processes also have lower ammonium sulfate usage due to having fewer precipitation stages, in the case of the second process simulation, and due to replacing precipitation wit ultrafiltration in the third process. The fourth process showed a higher C-PC purity of > 4.00, compared to the 3.77 simulated for the original process, and a C-PC recovery of 42.7 % from the crude extract. The chemical consumption was similar to the original process, while decreased amounts of ammonium sulfate were required. This process showed the shortest path time of the five options. Of the process configurations presented, the fifth one is best recommended, since it leads to a short batch-time (24.8 h per 3 batches), fewer process units than the original and the lowest overall chemical usage of the five processes. It also produces higher C-PC purification and recovery with reduced complexity compared to the original process. A C-PC purity of up to or above 4.0 is estimated to be achievable with this process, at a recovery of 47.7 % (using the leached C-PC as the starting point). This process produces a C-PC quality well above cosmetic grade. The trade-off between recovery and purity could be explored to achieve a higher recovery of cosmetic grade C-PC. To summarise, phase diagrams were produced for the PEG – MDX ATPS, and the C-PC leaching was tested on the Spirulina used in this project. This ATPS was then tested before moving on to a two-stage ATPS, using a PEG – citrate stage, for which PEG – citrate phase diagrams were produced, before the PEG – MDX stage. This produced better results, comparable to PEG – salt ATPS studies found in literature. Results from the process simulations done on Superpro Designer then supported the use of a pre-treatment step before a single-phase PEG – MDX ATPS, followed by precipitation and spray drying, based on information from this study and other literature. This could lead to a feasible design for pilot-testing and a novel process for industrial C-PC production. It is recommended that the results of the simulations be tested experimentally in further studies. A thorough techno-economic analysis of the proposed processes is also required. The pre-treatment process based on adsorption requires experimental validation and pilot scale confirmation before being applied in industry. Due to the rapidly growing demand for C-PC, a commercial process capable of producing multiple grades of C-PC, from foodgrade, to cosmetic-and reagent-grade, could be lucrative for business interests involved.
- ItemOpen AccessInvestigating process stresses on Saccharomyces cerevisiae using isothermal microcalorimetry(2017) Myers, Matthew; Harrison, Susan TL; Tai, Siew; Huddy, Robert; Fagan-Endres, MarijkeMaximising performance of microbial processes, including yeast-based processes, in an industrial setting requires understanding of the impact of process stresses. These may be the result of process configuration, dilution, temperature changes, hydrodynamic conditions or process perturbations. Methods to determine the microbial metabolic response to such stresses have long been sought, but are typically limited, often requiring the use of a suite of methods to assess the physiological status and state. The recent technical advances in microcalorimetry suggest potential for the use of isothermal microcalorimetry (IMC) to determine yeast viability and vitality and is investigated here. IMC is a laboratory method whereby the real-time heat produced by a chemical, biological or physical process is measured in the micro to nano watt range. It is proposed that this heat production may be correlated to the physiological state of the microbial catalyst and can be used to measure the impact of different stresses. In this study, the potential of IMC as a method for exploring process stress is investigated using Saccharomyces cerevisiae and its application in the beer brewing industry as a case study. Here, it is well known that yeast viability and vitality have commercial significance. IMC is sufficiently sensitive to detect the heat given off by 1000 yeast cells. However, IMC cannot distinguish between different heat flows within a system i.e. it is non-specific. The literature demonstrates how IMC has been used in the study of numerous microbiological fields, including the growth and metabolism of yeast. Previous studies have successfully derived the specific growth rate and cell numbers of a growing yeast population from analysing power and heat curves. The specific growth activity and specific growth retardation of yeast and how these parameters relate to bactericidal and bacteriostatic effects has also been examined by a number of authors. The key objectives of this study were to determine the viability and vitality of Saccharomyces cerevisiae using IMC and to assess the impact of stresses on yeast viability and vitality. This was achieved by measuring the thermal power produced by a growing yeast suspension as a function of its overall growth and metabolism. Two industrially relevant stresses were examined: cold shock and ethanol shock. The effect of these stresses has yet to be studied using microcalorimetry. The growth of Saccharomyces cerevisiae under ethanol stress was used as an inhibition study to isolate its effects on the growth thermogram. Following the generation of thermograms under control and stress conditions using IMC, a method for their quantitative analysis was developed. Curves were fitted to the heat data using an exponential growth equation and the time for the heat flow curve to peak was determined. From the exponential curve, the specific growth rate of the yeast was determined with a high degree of repeatability. The coefficient of the exponential term in the growth equation gave highly reproducible and distinguishable results relating to the viability and vitality of the initial yeast population. The time of peak heat flow was also affected by the initial viability and vitality of the yeast and was used to estimate the initial active cell population size.
- ItemOpen AccessInvestigating variables affecting heap (bio)leaching through determining access to sub-surface mineral grains by micro-scale X-ray tomographyGhadiri, Mahdi; Fagan-Endres, Marijke; Harrison, SusanHeap bioleaching is a hydrometallurgical technology, used to facilitate the extraction of valuable metals such as copper, gold, nickel and uranium from low-grade, typically sulphidic, ores. The process is highly complex as it is influenced by interactions of different sub-processes including flow of leaching solution around the ore particles, mass and heat transfer within and around the particles, chemical reactions, microbially-mediated reactions and microbial growth. Contact of leaching solution with mineral grains is necessary for oxidation of the sulphide minerals. However, a large fraction of the mineral grains is positioned below the surface of the ore particles and so contact with the liquid occurs through cracks and pores in the ore connected to the surface. Long extraction times and low metal recoveries typical of heap systems can be attributed to the slow leaching rate of these non-surface mineral grains as well as constraints on their accessibility. Most of the valuable grains that remain in the residue ores are non-surface grains. Therefore, investigation of the mechanism and behaviour of non-surface grain leaching and quantification of the factors contributing to their leaching is expected to be highly beneficial in the optimisation of leach conditions and recoveries. Non-surface grain leaching within large particles cannot be investigated via traditional experimental methods reliant on bulk measurements, 2D or destructive methodologies. However, it can be studied using high resolution, non-destructive 3D X-ray micro-Computed Tomography (μCT), an imaging technique for investigation of internal structure of opaque objects. X-ray μCT has previously been developed and used for investigation of different aspects of heap leaching. In the current study, the viability of using X-ray μCT to study heap bioleaching systems and affecting variables is assessed. This required establishment of procedures for measurement and analysis of sulphide and oxide mineral recoveries and leaching penetration distances. The feasibility of studying biotic heap leaching by X-ray μCT was explored through investigation of the relative energies required for high mineral resolution and avoidance of microbial inactivation. Specific bioleaching operating variables that were subsequently considered included: the accuracy and representivity of the X-ray μCT images, the influence of agglomeration pre-treatment, operating temperature, and type of ore on non-surface grain leaching. Addition of surfactants to the leaching solution was explored with the aim of changing surface activity to influence the penetration of the leach agent into pores and cracks in the ore. The effects of operating conditions on non-surface mineral grain leaching was studied using mini-column experiments. Three different low-grade ores, namely a chalcopyrite-rich ore, a malachite ore and a waste rock containing pyrite were prepared for the leaching experiment. The ores were crushed using a jaw crusher and comminuted down to 100% passing 16 mm. The products were sieved into six fractions (<0.25 mm, 0.25 - 1 mm, 1 - 2 mm, 2 - 5.6 mm, 5.6 - 8 mm, 8 - 16 mm) and each fraction then representatively split into smaller portions using a rotary splitter. One portion of each size fraction was taken for XRD, AAS and QEMSCAN analyses. Mini leaching columns were designed and constructed based on the target mineral grain distribution in the ores to ensure that the mineral grains were detectable using X-ray µCT, given its resolution limitations. The columns were charged with 50 g of agglomerated or non-agglomerated ore and lixiviant was provided at a flow rate of 2.55 mL h -1 for a period of 5.5 months for chalcopyrite and pyrite and 26 days for malachite in incubators at 30 °C, 37 °C and 65 °C. In order to select a surfactant suitable for use in a biological leach experiment, the effect of five different types and concentration of non-ionic surfactants on bioleaching microorganisms was studied in terms of microbial growth, ability for ferrous ion oxidation and chalcopyrite bioleaching. This was done in shake flask experiments using mineral concentrate. Based on the results of these experiments, Tween® 20 (10 mg L -1 ) was selected to study the effect of surfactant on non-surface mineral grain leaching in the mini-columns. Each column was scanned by X-ray μCT at 100 kV and 150 mA using a 0.38 mm copper filter and at a distance of 59.40 mm between X-ray gun and specimen. The advanced 3D analysis software Avizo® 9 was used to visualize and analyse image data. The Interactive Thresholding function in Avizo® 9 software was used for segmentation of ore particles from air and sulphide minerals from air and gangue minerals, to measure the target minerals' volume reduction during leaching. The Distance Map Algorithm was applied on a binary (segmented) image to calculate the distance of the sulphide mineral from the ore particle surface. Imaging of the whole mini-column was done before leaching and at the end of each experiment and imaging of certain sections was done at select time points during leaching to track temporal leaching dynamics. Good agreement was seen between the bulk mineral recovery data, determined using standard chemical assays, and the leaching curves generated using the X-ray µCT images for all the ores, confirming that the X-ray µCT images were a good quantitative measurement of the sulphide and oxide mineral leaching. Liquid microbial culture experiments were used to confirm that exposure to X-ray does not affect microbial activity for energy doses between 35 and 90 kV at 200-280 μA. However, X-ray exposure was found to have a slight negative influence at higher voltages of 120 and 150 kV, temporarily reducing the specific ferrous ion oxidation and suppressing the specific growth rate of the bioleaching microorganisms. The X-ray exposure thus negatively affected both the total microbial population available for leaching (population viability) as well as the metabolic activity of the individual microorganisms (population vitality). The effect of X-ray exposure on bioleaching cultures attached to a mineral surface was examined using pyrite-coated glass beads packed into mini-columns. The energy dosage limits identified in the liquid culture experiments were found to be compatible with the X-ray μCT imaging conditions (minimum energy dosage and sample position) required for acquisition of complete and accurate images of the columns at a resolution that allows identification of individual mineral grains. Following X-ray exposure, the performance of the exposed bioleaching mini-columns was equivalent to the unexposed control column. Similarly, the microbial activity and presence on the mineral surface appeared unchanged. Finally, the experiment was performed on the chalcopyrite ore and the microorganisms were found to still be able to convert Fe2+ to Fe3+ after 2 scanning runs. Thus, all sets of results confirm that X-ray μCT can be compatible with heap bioleaching experiments, while still permitting appropriate resolution of the mineral grains to make an X-ray μCT investigation worthwhile. However, cognisance that an upper limit of tolerable X-ray exposure exists must be taken. This may present a challenge if it is desired to image larger or denser ore samples which require a greater X-ray energy level for sufficient penetration of the sample by the X-rays and hence accurate imaging. In chalcopyrite leaching, increasing temperature from 37 °C to 65 °C resulted in clear enhancement of leaching based on both analysis methods, with the copper recovery increasing from 20% to 64% by the end of the leaching period, and the overall sulphide mineral dissolution increasing from 24% to 67%. Increasing temperature from 37 °C to 65 °C resulted in an increased leaching penetration distance and crack development in the particles, and thus an enhancement in copper recovery and sulphide mineral dissolution. This was in addition to the thermodynamically expected increased leaching rate. The maximum leaching penetration distance, beyond which no mineral volume change is observed, at 37 °C was 1.7 mm. This increased to 2.5 mm at 65 °C. As a result of addition of 10 mg L-1 Tween® 20 into the leaching solution, the final copper recovery was improved by 4% to 68% and the maximum penetration distance increased to 2.9 mm. However, when the availability of sulphide mineral was not rate limiting, the copper recovery and sulphide mineral volume reduction in the mini-column with surfactant was lower than the system without surfactant. This may have been due to depression of diffusion of ferric ion to the ore surface as a result of the formation of an adsorbed surfactant layer on the mineral surface. The performance with surfactant became superior as the amount of readily leachable mineral became limiting. In the pyrite waste rock, an increase in temperature did not have any effect on the maximum penetration distance and any increase in iron recovery was only for thermodynamic reasons. Similarly to the chalcopyrite ore, during the later period of leaching when readily exposed mineral grains have been depleted, the system performed better in the presence of surfactant. The addition of surfactant increased the maximum penetration distance from 2.7 to 2.9 mm. The cumulative copper recovery of 86% was obtained for malachite ore in 26 days of acid leaching and the maximum penetration distance was 2.2 mm. This study thus demonstrates the value of the X-ray µCT technique for quantitative investigation of non-surface mineral grain leaching and confirms that the maximum penetration distance can be affected with changing operation conditions or ore type. This study thus demonstrates the X-ray µCT technique for quantitative investigation of non-surface mineral grain bioleaching and confirms that the maximum penetration distance can be affected with changing operation conditions. Critically, the results confirm that X-ray μCT can be compatible with bioleaching microorganisms, while still permitting appropriate resolution of the mineral grains to make an X-ray μCT investigation worthwhile.
- ItemOpen AccessMagnetic resonance imaging characterisation of the influence of flowrate on liquid distribution in drip irrigated heap leaching(Elsevier, 2015-12) Fagan-Endres, Marijke; Harrison, Sue; Johns, Michael L; Sederman, Andrew JLiquid irrigation is one of the key process control parameters following the construction of an ore leaching heap. This study uses 3D magnetic resonance imaging (MRI) to examine non-invasively the effect of liquid flowrate changes on heap hydrology when drip irrigation is used. Experimental results from a vertical column show that the increase in flowrate causes an increase in the number of rivulets in the ore bed. The new rivulets were found to be thicker, and their development caused an increase in liquid-solid contacting area which is considered advantageous for metal ion recovery. Experiments performed on larger samples showed that the effects of flowrate changes were limited to the region directly below the drip emitter because the increase in flowrate caused an increase in macro-pore flow and not capillary retention of liquid. Therefore the increase in flowrate was not found to perturb liquid distribution patterns in a way that would be substantially advantageous to heap leaching recoveries.