3. Metodología y datos
3.2. Descripción de la metodología PCA
The waste rock was pre-washed for 24 hours prior to being inoculated to remove acid soluble material. Following inoculation, the pH of the un-inoculated PEL-LS control in the suspended mineral batch biokinetic test (BT) increased to pH 2.77 on day 4 (Figure 7.2 A), whereafter it remained constant till day 13, before declining to reach pH 2.1 on day 23. The un-inoculated PEL-HS reached a maximum pH of 3.48 on day 6, before it declined to pH 1.97 on day 23 (Figure 7.2 A). Despite these samples being not inoculated, the decline in pH could be attributed to the action of indigenous iron- and sulfur oxidising microorganisms inherent to the waste material as this material had not been sterilised. The pH of the inoculated PEL-LS sample increased to pH 2.18 on day 4 and declined to pH 1.51 by day 12 (Figure 7.2 A), whereafter it remained relatively constant. The pH of inoculated PEL-HS sample also reached a maximum of pH 2.2 on day 4, then declined to 1.51 by day 11 (Figure 7.2 A). The decline in pH is attributed to activity of the inoculated microbial culture and the indigenous microbes facilitating the degradation of the waste rock surface.
In the batch biokinetic test using coated beads (BT-CB), the waste rock mineral coated beads were sterilised prior to the experimental run. The pH of un-inoculated PEL-LS increased to pH 2.97 on day 17 (Figure 7.2 B) and the un-inoculated PEL-HS increased gradually to maximum pH of 3.48 on day 21 (Figure 7.2 B), before decreasing slightly. The extended acid neutralisation resulted from the absence of onset of acidification owing to the absence of iron- and sulfur-oxidising microorganisms. The pH of inoculated PEL-LS test declined from a maximum of pH 1.75 on day 3 to pH 1.45 on day 30 (Figure 7.2 B) and the pH of PEL-HS test
declined from pH 1.78 on day 3, to 1.28 on day 18 (Figure 7.2 B). Waste rock mineral-coated beads packed into the column reactors (FT-CB) and irrigated at 60 ml h-1 flow rate, were also
sterilised prior to experimental run, and unlike the batch shake flasks, were operated continuously in an open circuit with a feed containing 0.5 g L -1 Fe2+ as FeSO
4 7H2O, at pH 1.6
(Figure 7.2 C). Following inoculation during which pH decreased, the pH of the samples increased from pH 1.38 to 1.77 for PEL-HS (Figure 7.2 C) and 1.6 for PEL-LS (Figure 7.2 C). The pH in the un-inoculated samples increased from pH1.6 to 2.01 for PEL-HS (Figure 7.2 C) and 1.69 for PEL-LS (Figure 7.2 C). The pH increase in the effluent suggests an initial dissolution of acid neutralising minerals such as calcite. During the complete flow-through operation phase, the average pH for the experimental PEL-HS and PEL-LS samples were 1.61 (± 0.035) and 1.59 (± 0.011) respectively and the average pH for control samples was 1.63 (± 0.024) for PEL-HS and 1.61 (± 0.022) for PEL-LS. These remained relatively stable and similar to the pH 1.6 media fed to the column reactors continuously in flow-through operation. This suggests that the residence time in the column was too low to liberate these compounds significantly to be detected by solution measuring techniques. The relatively constant pH also suggests a lack of detectable activity in the leaching of the available sulfidic mineral in both waste rocks. It was shown in Chapter 6 that despite having a lack of detection of leaching activity in solution chemistry at 60 ml h-1, there was evidence of leaching activity
when mineral colonising microbial populations were measured directly.
In the waste rock mineral-coated beads packed into the column reactors (FT-CB) at 4 ml h-1
flow rate, the pH of un-inoculated column containing PEL-LS increased to pH 2.83 on day 2 due to acid neutralising capacity being solubilised, before declining to pH 1.6 by day 12 and a lowest point of pH 1.44 on day 20 (Figure 7.2 D) due to acidification and the wash out of acid neutralisation. In the PEL-HS column, the pH increased to pH 3.2 on day 2, before declining to pH 1.65 by day 12 and a minimum of pH 1.54 on day 21 (Figure 7.2 D). The decline in pH could attributed to the depletion of neutralising minerals, removed by the flow-through nature of the column. The pH in the inoculated PEL-LS increased only to pH 2.27 on day 4 owing to the early onset of microbially-mediated acidification, thereafter decreasing to pH 1.45 by day 11 and to pH 1.37 on day 20 (Figure 7.2 D). In the PEL-HS column, the pH increased to pH 2.25 on day 4, before declining through pH 1.5 on day 8 and pH 1.25 on day 20 (Figure 7.2 D). The pH profiles of the inoculated tests across the three biokinetic tests (Figure 7.2 A, B and D) showed that microbial activity facilitated leaching of the sulfidic waste rock and that the rates of acidification and neutralisation were not equivalent. The pH profiles of the flow- through system at 60 ml h-1 flow rate showed a lack of significant detectable change between
Didi X Makaula Chapter 7
effectively detect activity in the solution during leaching of the available sulfidic mineral in both waste rocks.
Figure 7.2: Analysis of pH across the biokinetic test approaches, (A) batch slurry (BT), (B) batch waste rock coated glass beads (BT-CB), (C) waste rock coated glass beads in a flow-through column (FT-CB; 60 ml h-1) and (D) waste rock coated glass beads in a flow-through column (FT-CB; 4 ml h-1). All experiments were conducted at 30 °C. The samples include PEL-LS un-inoculated (), PEL-HS un-inoculated (), PEL-LS inoculated (◆) and PEL-HS inoculated (). Error bars represent the standard deviation from the mean pH across the three experimental shake flasks and five columns at the same time point.
Didi X Makaula Chapter 7
In the batch biokinetic test with un-sterilised suspended mineral, the redox potential of both the un-inoculated PEL-LS and PEL-HS remained in the range ~250 mV to 300 mV until day 10. Thereafter, it increased gradually to a maximum of 666 mV and 649 mV by day 22 in the PEL-LS (Figure 7.3 A) and PEL-HS tests (Figure 7.3 A) respectively. The redox potential in the inoculated PEL-LS increased from 480 mV at the start of the run to 712 mV on day 4 and remained relatively stable thereafter (Figure 7.3 A); similarly, the PEL-HS test redox potential increased from 468 mV to 696 mV in the same period and then stabilised (Figure 7.3 A). In the BT-CB using sterilised mineral, the redox potential of un-inoculated PEL-LS (Figure 7.3 B) and PEL-HS (Figure 7.3 B) tests remained in the range 250 to 300 mV over the 17 and 25 day duration of the runs respectively. This can be attributed to the absence of indigenous microbial populations due to sterilisation.
In the inoculated BT-CB tests, the redox potential of PEL-LS increased from 498 mV on day 1 to 683 mV on day 7 (Figure 7.3 B) and that of PEL-HS from 488 mV on day 1 to 710 mV on day 10 (Figure 7.3 B) and remained relatively stable throughout the experimental run. In the flow-through tests using sterile mineral (FT-CB) at 60 ml h-1 flow rate, the redox potential of
the inoculated samples decreased from 499 to 401 mV for PEL-HS (Figure 7.3 C) and from 504 to 430 mV for PEL-LS during the 18-hour inoculation period. The redox potential of the un-inoculated samples remained the same at 301 mV for PEL-HS (Figure 7.3 C) and increased from 301 to 335 mV for PEL-LS. The redox potential of both PEL-HS and PEL-LS inoculated samples remained relatively low, circa 300 to 500 mV, throughout the continuous flow-through operation (Figure 7.3 C). This suggests either a lack of effective and detectable iron-oxidizing microbial activity that would result in the catalysed regeneration of Fe3+ and
associated increased redox potential, or that the microbial activity generating Fe3+ is
insignificant with respect to the Fe3+ leaching of the mineral and the flow rate of the solution.
When this is related back to the pyrite concentrate study (Chapter 6) that was operated at the same flow rate (60 ml h-1), a high redox potential was recorded reaching a maximum of 696
mV (Figure 6.3). This suggests that the mineral grade plays a role in determining microbial- mineral interaction and thus the facilitation of leaching.
In the un-inoculated PEL-LS and PEL-HS flow-through tests using sterile mineral (FT-CB) at 4 ml h-1 flow rate, the redox potential remained low throughout the 30 day experimental run,
increasing gradually from 300 to 400 mV (Figure 7.3 D). An increase in redox potential of the inoculated PEL-LS test was evident from day 10 (391 mV) to day 21 (700 mV; Figure 7.3 D), whereafter it remained relatively stable. The redox potential of inoculated PEL-HS test increased from 416 mV to 689 mV in the same period (Figure 7.3 D). Based on solution chemistry, both waste rock minerals are acid forming across three biokinetic systems (Figure 7.3 A, B and D), supporting the metabolism of iron- and sulfur-oxidising microorganisms. At
60 ml h-1 flow rate, not enough evidence is provided in the solution chemistry, both pH and
redox potential, to suggest that in the inoculated samples there is active degradation of both waste rocks facilitated by microbial populations (Figure 7.3 C)
Didi X Makaula Chapter 7
Figure 7.3: Measured redox potential across the biokinetic test approaches, (A) batch slurry (BT), (B) batch waste rock coated glass beads (BT-CB), (C) waste rock coated glass beads in a flow-through column (FT-CB; 60 ml h-1) and (D) waste rock coated glass beads in a flow-through column (FT-CB; 4 ml h-1). All experiments were conducted at 30 °C. The samples include PEL-LS un-inoculated (), PEL-HS un-inoculated (), PEL-LS inoculated (◆) and PEL-HS inoculated (). Error bars represent the standard deviation from the mean redox potential across the three experimental shake flasks and five columns at the same time point
In the flow-through biokinetic system at 4 ml h-1 flow rate, it has been shown through both
solution pH and redox potential profiles that the inoculated microbial populations facilitated the degradation of the two waste rock minerals, albeit requiring longer to establish the leaching environment. Moreover, acid neutralising capacity was washed out of the columns during the early onset of acidification, negating its role in neutralisation. The 0.5 g L-1 Fe2+ supplemented
at the beginning of the experimental run, as well as that generated through mineral leaching in the batch biokinetic tests (Figure 7.4 A and B) , and the 0.5 g L-1 Fe2+ that was continuously
fed in flow-through biokinetic system at 4 ml h-1 (Figure 7.4 D), was dominantly oxidised across
three inoculated waste rock samples. The same amount of Fe2+ supplemented in the flow-
through biokinetic system at 60 ml h-1, remained relatively the same throughout when
measured in the effluent. By day two, the Fe2+ was completely depleted by the microbial
populations present in both inoculated PEL- LS and PEL-HS respectively (Figure 7.4 A) and this was maintained throughout the experimental run. In the absence of oxidising microbial populations, the 0.5 g L-1 Fe2+ supplemented in the un-inoculated increased gradually and
reached a cumulative maximum of 1.36 g L-1 for PEL-LS on day 10 (Figure 7.4 A) and 1.53 g
L-1 for PEL-HS on day 12 (Figure 7.4 A), thereafter oxidation was evident, suggesting the
presence of indigenous oxidising microbial populations. In the BT-CB test, depletion of Fe2+
was achieved by day 5 for both PEL-LS and PEL-HS respectively (Figure 7.4 B). In the sterilised and un-inoculated samples, the Fe2+ increased gradually and reached a cumulative
maximum of 1.21 g L-1 on day 12 for PEL-LS (Figure 7.4 B) and 1.31 g L-1 on day 19 for PEL-
HS (Figure 7.4 B).
In the FT-CB system at 60 ml h-1 flow rate, the Fe2+ decreased from 0.841 at day 0 to 0.382
g L-1 at day for PEL-LS (Figure 7.4 C) and 0.831 at day 0 to 0.495 g L-1 at day 2 for PEL-HS
(Figure 7.4 C). The measured Fe2+ remained relatively similar to the fed 0.5 g L-1 throughout
the experimental run. The measured effluent of Fe2+ in the inoculated PEL-LS (Figure 7.4 C)
and PEL-HS (Figure 7.4 C) samples also remained relatively similar to the fed Fe2+. Fe2+ was
largely oxidised into Fe3+ under the same flow rate on a pyrite concentrate (Figure 6.4; Chapter
6). At a lower flow rate of 4 ml h-1 (Figure 7.4 D), the Fe2+ of the sterile and un-inoculated
increased from 0.5 at day 0 to 1.11 g L-1 at day 1 for PEL-LS (Figure 7.4 D) and increased
from 0.5 at day 0 to 1.14 g L-1 at day 1 for PEL-HS (Figure 7.4 D). The initial increase in Fe2+
in both samples suggested that there was a washout of readily leachable Fe2+ on the waste
rock surfaces and was also due to the absence of oxidising microbial populations. The Fe2+
decreased gradually until a concentration similar to the feed was achieved and then it remained relatively stable throughout the experimental run. In the inoculated samples, the Fe2+ reached a maximum of 1.87 g L-1 for PEL-LS (Figure 7.4 D) and 1.67 g L-1 for PEL-HS
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Figure 7.4: Measured Fe2+ across the three biokinetic test approaches, (A) batch slurry (BT), (B) batch waste rock coated glass beads (BT-CB), (C) waste rock coated glass beads in a flow-through column (FT-CB; 60 ml h-1) and (D) waste rock coated glass beads in a flow-through column (FT-CB; 4 ml h-1). All experiments were conducted at 30 °C. The samples include PEL-LS un-inoculated (), PEL-HS un-inoculated (), PEL-LS inoculated (◆) and PEL-HS inoculated (). Error bars represent the standard deviation from the mean Fe2+ across the three experimental shake flasks and five columns at the same time point.
The Fe2+ decreased and was depleted by day 15 in both waste rock samples due to microbial
activity facilitating its oxidation and Fe2+ remained oxidised throughout rest of the experimental
run. The Fe3+ trend across the biokinetic tests (Figure 7.5 A, B, C, and D) corresponded with
the trends of Fe2+ (Figure 7.4) and redox potential (Figure 7.3) counterparts. The Fe3+ of the
un-inoculated waste rock samples increased from 0.06 on day 13 to 1.31 g L-1 on day 23
(Figure 7.5 A) and from 0.19 on day 13 to 1.53 g L-1 on day 23 (Figure 7.5 A). The increase
suggested that there was a presence of oxidising microbial populations that facilitated the regeneration of Fe3+ and this corresponded with the decline in Fe2+ (Figure 7.4 A and B) and
an increase in redox potential (Figure 7.3 A and B). The Fe3+ in the un-inoculated and sterilised
samples for both PEL-LS and PEL-HS remained relatively low with now indication of any accelerated oxidation (Figure 7.5 B, C and D). The Fe3+ of the inoculated increased gradually
from 2.29 to 3.6 g L-1 for PEL-LS and from 2.34 to 4.1 g L-1 for PEL-HS over 23 days (Figure
7.5 A). The Fe3+ increased from 1.25 to 4.58 g L-1 for PEL-LS and from 1.33 to 7.4 g L-1 for
PEL-HS over 30 days (Figure 7.5 B). The Fe3+ remained relatively unchanged for both PEL-
LS and PEL-HS throughout the continuous flow-through experimental run at 60 ml h-1 flow rate
(Figure 7.6 C). Conversely, at 4 ml h-1 flow rate, an increase in Fe3+ between day 14 and 16
was observed in both inoculated PEL-LS and PEL-HS and thereafter the measured effluent Fe3+ was in excess of the fed and oxidised Fe2+, suggesting that there was active degradation
of the sulfidic waste rock surface facilitated by oxidising microbial populations (Figure 7.5 D). Solution chemistry analysis between the two waste rocks at the two different flow rates generally showed completely different outcomes. At high flow rates, changes in solution chemistry were not detectable in the effluent, whereas at lower flow rates performance of the system (both pyrite sulfidic waste rocks) was detected and noticeable. It is also worth noting that in Chapter 6, when the same high flow rates were applied in a pyrite concentrate system, the changes in solution chemistry were detectable. This suggests that the grade of a mineral had a significant impact on microbial behaviour and performance
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Figure 7.5: Measured Fe3+ across the three biokinetic test approaches, (A) batch slurry (BT), (B) batch waste rock coated glass beads (BT-CB), (C) waste rock coated glass beads in a flow-through column (FT-CB; 60 ml h-1) and (D) waste rock coated glass beads in a flow-through column (FT-CB; 4 ml h-1). All experiments were conducted at 30 °C. The samples include PEL-LS un-inoculated (), PEL-HS un-inoculated (), PEL-LS inoculated (◆) and PEL-HS inoculated (). Error bars represent the standard deviation from the mean redox Fe3+ across the three experimental shake flasks and five columns at the same time point