Environ Geochem Health
DOI 10.1007/s10653-007-9095-z
ORIGINAL PAPER
The effects of Acidithiobacillus ferrooxidans on the leaching
of cobalt and strontium adsorbed onto soil particles
Hyun-Sung Park Æ Jong-Un Lee Æ Jae-Woo Ahn
Ó Springer Science+Business Media B.V. 2007
Abstract Bioleaching from soil artificially contam-
inated with analogues of radionuclides, Co and Sr,
was carried out using a Fe-oxidizing bacterium,
Acidithiobacillus ferrooxidans. Due to bacterial
metabolism, the pH and dissolved Fe3+ concentration
in a biotic slurry decreased and increased respec-
tively, over time, but the concentrations of Co and Sr
extracted from the soil showed no significant
enhancement compared with those under abiotic
control. In both cases, Co and Sr were leached from
the soil during the initial period of the experiment,
due to the initially low solution pH of 2.0, and the
dissolved concentrations remained almost constant
for the duration of the experiment (300 h). Since
oxidation of Fe2+ by A. ferrooxidans led to the
production of Fe precipitates and colloidal suspen-
sions, the Co and Sr extracted into solution were most
likely re-adsorbed onto the Fe solids. Also, A.
ferrooxidans, without an external supply of Fe2+,
extracted almost equal or greater amounts of Co and
Sr from the soil than when Fe2+ was supplied. Under
H.-S. Park
J.-U. Lee (&)
Microbial Geochemistry Lab, Department of Geosystem
Engineering, Chonnam National University, Gwangju
500-757, Korea
e-mail: jongun@chonnam.ac.kr
J.-W. Ahn
Department of Advanced Materials Science and
Engineering, Daejin University, Pocheon 487-711, Korea
the same leaching conditions, the extent of Sr
removal was much lower than that of Co. On the
contrary to the high efficiency of microbial metal
leaching in biohydrometallurgy for low-graded sul-
fide ores, which has been widely documented,
conventional bioleaching techniques with A. ferroox-
idans supplied with enough Fe2+ showed low effi-
ciency for the removal of radionuclides loosely bound
onto soil particle surfaces.
Keywords Bioleaching
Radionuclides
Cobalt

Strontium
Acidithiobacillus ferrooxidans
Soil
Introduction
Contamination of soil and sediment with radionuc-
lides can pose significant hazards to human health. In
particular, unexpected accidents at nuclear power
plants, e.g., in Chernobyl, and potential release of
radioactive elements from disposal sites, will signif-
icantly exert malign influences on the biosphere.
When surface environments are polluted with radio-
nuclides, the application of natural containment
technology, through sorption or precipitation, will
never be a radical remediation method due to the very
long half-lives of most radionuclides. Therefore, once
soil and sediment pollution with radioactive elements
has occurred, thorough clean-up processes should be
performed immediately.
123

Chemical and bacterial leaching methods have
been developed for the decontamination of soil and
sediment polluted with heavy metals. Chemical
treatment, with various oxidizing, reducing, and
complexing agents, has been used for the extraction
of heavy metals from contaminated soil, sediment,
and sewage sludge (e.g., Kedziorek et al. 1998; Steele
and Pichtel 1998; Wasay et al. 1998). However, the
high cost, operational difficulties, intensive energy
requirements, unsatisfactory yield of metal solubili-
zation and hazardous byproducts have caused
researchers to consider alternative methods (Tyagi
and Couillard 1989). The bioleaching process has
been proposed as an effective, economic, and envi-
ronmentally-friendly method for the treatment of
heavy metals and radionuclides.
One of the most widely used microorganisms in
bioleaching is the acidophilic Acidithiobacillus fer-
rooxidans (Blais et al. 1992). This lithoautotrophic
bacterium gains energy to support growth and
maintenance from the oxidation of Fe2+ or reduced
sulfur compounds, and uses CO2 as a cellular carbon
source. Therefore, an additional carbon source or
electron donor is not required. When A. ferrooxidans
oxidizes Fe2+ to Fe3+, it produces protons and an
oxidant (Fe3+), which are critical factors in the
leaching process, especially of sulfides.
Acidithiobacillus ferrooxidans oxidizes ferrous to
ferric ions (Eq. 1), with the latter subsequently
leading to the oxidative dissolution of sulfides (Eq.
2). Overall reactions produce extra hydrogen ions
and, thus, the system pH is lowered (Boon et al. 1998;
Fowler and Crundwell 1998).
bacteria
14 Fe2þ + 3.5 O2 + 14 Hþ ! 14 Fe3þ + 7H2 O
ð1Þ
þ contaminated soil, sediment, and sewage sludge using
MeS2 +8H2 O+14Fe3þ !Me2þ +14Fe2þ +2SO2
4 +16H
ð2Þ
(MeS2: insoluble metal sulfides, Me2+: free metal
ion)
Some researchers have suggested that the direct
contact of bacteria with sulfide surfaces results in
metal dissolution (Eq. 3; Silverman and Ehrlich 1964;
Free et al. 1991; Lizama and Suzuki 1991). However,
according to some recent well-designed investigations
123
Environ Geochem Health
(Schippers et al. 1996; Fowler and Crundwell 1998),
the bacterial production of Fe3+ and subsequent sulfide
dissolution play a dominant role in bioleaching
processes.
bacteria þ
MeS2 + H2 O + 3.5 O2 ! Me2þ + 2 SO2
4 + 2H
ð3Þ
The processes described above (Eqs. 1–3) require
sulfides to be oxidized, with the production of
hydrogen ions, which finally lowers the system pH.
However, natural soil, sediment, and sludge contam-
inated with heavy metals generally contain only small
amounts of sulfur and sulfides, with the exception of
those related to mining activity. In these cases,
however, the acidification of non-sulfide solids by
A. ferrooxidans can also be performed by Fe2+
oxidation (Eq. 4), followed by the precipitation of
ferric iron in the form of jarosite (Eq. 5) or ferric
hydroxide (Gu and Wong 2004).
bacteria
4 Fe2þ + O2 + 4 Hþ ! 4 Fe3þ + 2H2 O ð4Þ
3 Fe3þ + Meþ + 2 HSO
4 + 6 H2 O
ð5Þ
! MeFe3 ðSO4 Þ2 ðOHÞ6 + 8 Hþ
(Me+: free metal ion)
Acidithiobacillus ferrooxidans has been practically
used in the hydrometallurgy of low-graded sulfide
ores since Fe3+ can lead to the oxidative dissolution
of sulfides (e.g., Dew et al. 1998; Dastidar et al.
2000). Moreover, much recent research has indicated
that the bioleaching technique can be successfully
applied to solving environmental problems related to
heavy metals. They reported many good results on
the effective biolixiviation of heavy metals from
Fe-oxidizing bacteria (e.g., Sreekrishnan et al. 1993;
Gomez and Bosecker 1999; Chen and Lin 2000; Jang
et al. 2002; Kim et al. 2005). The efficiency of the
microbial leaching process is affected by various
parameters, such as the bacterial strain used, the
quantity and type of substrate, and the solid content,
particle size, and carbon dioxide and oxygen con-
centrations in the system, as well as the pH and redox
potential (Couillard and Zhu 1992; Haddadin et al.
1995).
Environ Geochem Health
Despite extensive research on heavy metals biole-
aching, few studies have been performed on the
remediation of soil and sediment contaminated with
radionuclides. Since the released radioactive ele-
ments will pollute soil and sediment, mainly by
adsorption via air and natural waters, they will exist
in a loosely bound mode in geological media. Thus,
the contaminants in radioactively polluted soil and
sediment differ in their mode of occurrence from that
of sulfide ore minerals, where the metals reside in a
strong crystal lattice structure. Therefore, whether Fe-
oxidizing bacteria can leach the metals or radionuc-
lides adsorbed onto solid surfaces, with a high
leaching efficiency, remains to be determined.
In order for A. ferrooxidans to maintain active
metabolism during the bioleaching process, an exter-
nal supply of both Fe2+ and inorganic nutrients, such
as NH+4 , K+, HPO2 2+ 2+
4 , Mg , and Ca , is required.
Recently, Lee et al. (2005) reported that a supply of
inorganic nutrients from natural soil for A. ferroox-
idans led to a slightly higher leaching efficiency of
uranium than with the artificial, external addition of
nutrients. Since the cost of supplying nutrients to the
bacterial cells can be reduced, if this were the case,
the experimental determination of the optimal leach-
ing conditions will be required.
The objectives of this study were to examine the
bioleaching capacity of A. ferrooxidans for the
extraction of radionuclides adsorbed onto the surfaces
of soil particles, and provide fundamental informa-
tion on the highly efficient and economic conditions
required for the leaching process.
Materials and methods
Artificial contamination of soil samples
Uncontaminated soil samples were taken in the
vicinity of Yeonggwang nuclear power plant, which
is located in the southwestern part of Korea. The
soil was dried at room temperature, and the physi-
cochemical properties of soil (cation exchange
capacity, soil pH, loss-on-ignition, and textural
analysis) determined. The soil sample (<0.18 mm
diameter) was soaked in a mixed solution of Co and
Sr (each 1,000 mg/l) for 24 h. Instead of radioactive
60
Co (half-life = 5.27 years) and 90 Sr (half-
life = 28.8 years), non-radioactive 59CoCl2
6H2O
(Sigma-Aldrich) and 88Sr(NO3)2 (Sigma-Aldrich)
respectively were used to contaminate the clean soil.
After pollution, the soil sample was washed three
times with distilled, de-ionized water and dried at
358C. The concentrations of Co, Sr, and Fe in the
original, uncontaminated and the artificially contam-
inated soils were determined after extraction by aqua
regia (HCl:HNO3 = 3:1) for 1 h at 708C .
Bacteria and culture conditions
A freeze-dried culture of A. ferrooxidans (KCTC
2677) was obtained from the Korean Collection of
Type Culture. Cells were cultivated in 100 ml of 9 K
medium in a 500-ml flask (Silverman and Lundgren
1959) placed in a shaking incubator at 308C and
200 rpm. The 9 K medium consisted of two types of
solutions; solution A: 3 g (NH4)2
SO4, 0.1 g KCl,
0.5 g K2HPO4, 0.5 g MgSO4
7H2O, and 0.014 g
Ca(NO3)2
4H2O in 700 ml of distilled water, and
solution B: 45 g FeSO4
7H2O in 300 ml of distilled
water. Solutions A and B provided inorganic nutri-
ents and an energy source to the bacterial cells
respectively. Serial transfer was performed several
times to enhance the cell activity.
Leaching experiments with and without bacteria
Batch-type bioleaching and abiotic control leaching
experiments were conducted according to the fol-
lowing conditions:
– Experimental set (1): Soil (25 g) + solution A
(350 ml) + solution B (150 ml) + A. ferrooxidans
(50 ml)
– Experimental set (2): Soil (25 g) + solution A
(350 ml) + solution B (150 ml) + sterile distilled
water (50 ml)
The experiments were performed in 1,000-ml
conical flasks, each containing 25 g of artificially
contaminated soil and 500 ml of 9 K medium, in a
shaking incubator at 308C and 200 rpm for 300 h.
Solution A of the 9 K medium was autoclaved, and
solution B was sterilized by passing through a 0.2-mm
filter. The bioleaching flasks were inoculated with
50 ml of A. ferrooxidans, which had been cultured for
30 h. The pH of the solution was adjusted to 2.0 with
5 M H2SO4 and 0.1 M NaOH. Periodically, aliquots
were taken from each flask, filtered through an
123

0.2-mm filter and the pH as well as dissolved Fe2+,
total Fe, Co, and Sr concentrations of the solution
were determined.
Bioleaching experiments under various conditions
To determine the experimental conditions leading to
high bioleaching efficiency, the experimental sets
were designed, as shown below, with the leaching
capacities then compared:
– Experimental set A: Soil (25 g) + solution A
(350 ml) + solution B (150 ml) + A. ferrooxidans
(50 ml)
– Experimental set B: Soil (25 g) + distilled water
(350 ml) + solution B (150 ml) + A. ferrooxidans
(50 ml)
– Experimental set C: Soil (25 g) + solution A
(350 ml) + distilled water (150 ml) + A. ferro-
oxidans (50 ml)
– Experimental set D: Soil (25 g) + distilled water
(500 ml) + A. ferrooxidans (50 ml)
Experimental set (A) was designed to examine the
bioleaching under 9 K medium conditions, which are
those most widely employed for A. ferrooxidans
activity. In the case of set B, solution A (the inorganic
nutrients for cellular growth and synthesis) was not
supplied to the slurry. In experimental set C, solution
B (Fe2+, which acts as an energy source for growth
and maintenance of A. ferrooxidans), was absent. In
experimental set D, neither solution A nor B was
added. A. ferrooxidans was infused to the system
after 30 h of incubation in the 9 K medium. The pH
of the solution was adjusted to 2.0 with 5 M H2SO4
and 0.1 M NaOH. After the periodic removal of
aliquots from the batch-type slurries, the solution pH
and dissolved Fe2+, total Fe, Co, and Sr concentra-
tions were measured after filtration through a 0.2-mm
filter.
After 300 h, the residual soils in each flask were
removed, dried at room temperature, and the Co, Sr,
and total Fe then extracted by aqua regia digestion
and determined.
Analytical techniques
The pH was measured using a glass electrode
connected to an Orion Model 720A pH meter. The
concentration of ferrous ion was determined by a
123
Environ Geochem Health
modified ferrozine method (Stookey 1970). For this, a
0.1-ml sample was rapidly mixed with 4.9 ml of 0.1%
ferrozine solution in 0.5% pH 7.0 ammonium acetate
buffer (adjusted with NaOH). The solution was
agitated, and then assayed using a UV-vis spectro-
photometer (UV-MINI-1240) at 562 nm. The con-
centration of ferric ion was calculated by subtracting
the measured Fe2+ concentration from that of the total
dissolved Fe.
The total Fe, Co, and Sr concentrations in the
slurries and residual soils were determined using
ICP-AES (Jobin Ybon JY70+).
Results and discussion
Characteristics of the soil sample
The studied soil was slightly acidic (pH = 5.0), and
had a silty loam texture (sand 2.3%, silt 64.9%, clay
32.8%). The cation exchange capacity was 30.0 meq/
100 g, with loss-on-ignition of 4.5%. The concentra-
tions of Co and Sr in the uncontaminated soil were 30
and 21 mg/kg respectively. The artificially contam-
inated soil had 373 and 393 mg/kg of Co and Sr
respectively. After soaking for only 24 h, the levels of
Co and Sr contamination in the soil samples were 10
times those of the uncontaminated soil. The concen-
tration of Fe in the soil was 1.60%.
Leaching efficiency of A. ferrooxidans and abiotic
control
Batch-type leaching experiments were conducted
under A. ferrooxidans (the set 1) and abiotic control
(the set 2) conditions. The bacteria provided with
solutions A and B of the 9 K medium as external
nutrient and energy sources produced a greater
amount of protons, which resulted in a lower solution
pH than that in the abiotic control (Fig. 1). This result
indicated that the iron oxidizer included in set 1
performed normal metabolism, where A. ferrooxidans
oxidized Fe2+ to Fe3+, with H+ supplied into the
solution by the chemical formation of Fe precipitates
(Eqs. 4, 5); however, the abiotic control showed a
gradual increase in the solution pH with time, as
hydrogen ions were chemically consumed during the
protonation and dissolution of solid particles. Since
the pH is apparently one of the factors significantly
Environ Geochem Health
2.6
2.4
2.2
pH
2.0
1.8 Set (1)
Set (2)
1.6
0 50 100 150 200 250 300
Times (hours)
Fig. 1 Variation in pH values in the leaching experiments with
A. ferrooxidans and the abiotic control over time. Set 1,
microbial leaching with A. ferrooxidans; set 2, abiotic control
mediating the rate and extent of metal leaching, the
decrease in the pH due to A. ferrooxidans would
imply that the use of this bacterium can lead to
favorable conditions for the leaching of metals from
various solids, and this strongly acidic environment
will be maintained over a relatively long period.
The amount of Fe2+, used as the energy source in
the 9 K medium in these experiments, was 9 g/l. At a
given time during the leaching process, the difference
between input Fe (9 g/l) and total dissolved Fe
contents are an indication of the amounts of Fe
precipitates and colloidal suspensions. Also, the
difference between total dissolved Fe and Fe2+ will
give the amount of dissolved Fe3+. Aqueous Fe3+
serves as a strong oxidant, which greatly facilitates
metal extraction from solid substrates, especially
sulfides (Eq. 2). The production of Fe precipitates,
such as jarosite, can also lower the solution pH, thus
enhancing metal leaching (Eq. 5). Therefore, the
microbial production of aqueous Fe3+ in solution is
essential for an effective leaching process.
The A. ferrooxidans in experimental set 1 oxidized
nearly all the Fe2+ and produced approximately 4 g/l
of Fe3+ within 110 h (Fig. 2). The concentration of
total dissolved Fe also decreased over time in set 1.
The reduced amount of total dissolved Fe (about 5 g/l
in set 1) was considered to have formed Fe precip-
itates and colloidal suspensions. During experiment
1, the formation of reddish brown precipitates and
colloidal suspensions was readily observed in the
flask employed. The transformation of Fe2+ into a
variety of Fe minerals, such as jarosite, schwertman-
nite, goethite, and ferrihydrite, during bioleaching
10
Dissolved Fe concentrations (g/L)
8 dissolved
3+
Fe
6
4
2 3+
dissolved Fe
0
0 50 100 150 200 250 300
Time (hours)
Fe(T) in set (1) Fe2+ in set (1)
Fe(T) in set (2) Fe2+ in set (2)
Fig. 2 Variations in aqueous total Fe (Fe(T)) and Fe2+
concentrations in the leaching experiments with A. ferrooxi-
dans over time
processes has been extensively documented by many
researchers (e.g., Tuovinen 2004). However, the
abiotic control showed no significant decrease in
the total Fe concentration, and by the end of the
experiment, more than 85% of the dissolved Fe
existed as Fe2+.
The amounts of Co and Sr extracted showed no
remarkable differences between the biotic samples
and abiotic controls (Fig. 3). Moreover, the concen-
trations of Co and Sr in experimental set 1 were lower
than in the abiotic controls for the duration of the
experiment.
This result was most likely due to the mode of
occurrence of Co and Sr in the soil, as well as the
mineralogical type of solid to be leached. In the
experiment, the soil was soaked in the Co and Sr
solution for 24 h to simulate radionuclide pollution
and, thus, that the Co and Sr might become loosely
bound onto the soil surface through adsorption. With
regard to the conditions, an initial pH of 2 was low
enough to extract the observed amounts of Co and Sr
in the abiotic control. This can be confirmed by the
amounts of dissolved Co and Sr abruptly extracted
during the initial period of experiments.
If the Co and Sr are associated with sulfide
minerals, the low pH and large amount of dissolved
Fe3+ in set 1, which were the characteristic results of
the microbial leaching process (Figs. 1, 2), might
significantly enhance the rate and extent of leaching
over time.
The formation of Fe precipitates and colloidal
suspensions following the oxidation of Fe2+ to Fe3+
123

Dissolved Co concentrations (mg/L)
30
25
20
15
10
5 Set (1)
0
0 50 100
Time (hours)
Dissolved Sr concentrations (mg/L)
30
25
20
15
10
5 slightly higher than that in set A, which was also
0
0 50 100
Time (hours)
Fig. 3 Variations in the dissolved Co and Sr concentrations in
the leaching experiments with A. ferrooxidans
may explain why experimental set 1 showed slightly
lower concentrations of dissolved Co and Sr than the
abiotic control. The Fe precipitates and colloidal
suspensions probably adsorbed the dissolved Co and
Sr in the solution and, thus, mediated the aqueous
concentrations of these metals. In experimental set 2,
where no bacteria were infused, the solution was
observed to be less stained than in set 1, indicating
the formation of many fewer Fe precipitates and
colloidal suspensions. The chemical oxidation of Fe2+
without microbial reactions is known to be slower
than that of A. ferrooxidans by 105–106 times (Lacey
and Lawson 1970).
The fact that more Co than Sr was extracted both
microbially and chemically, as shown in Fig. 3,
indicates that the extraction efficiency may depend on
the type of radionuclides. Conceivably, Sr may be
more readily adsorbed onto other solids or formed
precipitates or colloidal suspensions than Co. Since
this alkaline earth element, which easily forms stable
sulfate precipitate, has not been the focus of environ-
mental investigations, the result is relatively new.
123
Environ Geochem Health
The bioleaching process using Fe-oxidizing bac-
teria has been proved to be one of the most promising
methods for the extraction of heavy metals from
contaminated soil and sediment. However, the pres-
ent study has indicated that such elevated bioleaching
efficiency can be constrained by the mode of
occurrence of the heavy metals, as well as the
mineralogical characteristics of soil and sediment.
Set (2)
150 200 250 300 Bioleaching under various operational conditions
Batch-type bioleaching was performed using exper-
imental sets A, B, C, and D, as described above. The
bacteria provided with solution B (Fe2+) of the 9 K
medium as an energy source (experimental sets A and
B) produced the high amounts of protons, resulting in
lower solution pHs than in experimental sets C and D,
where no Fe2+ was externally supplied (Fig. 4).
Set (1)
Although the pH in experimental set B was
Set (2)
provided with inorganic nutrients, a dramatic pH
150 200 250 300 decrease was observed over time. Considering that
metabolically active A. ferrooxidans continuously
produces protons, the bacterium, when not supplied
with inorganic nutrients, also appeared to perform
active metabolism. As described earlier, this result
was probably due to the natural supply of a variety of
inorganic nutrients to the bacteria from the soil
substrate. The elements leached from the soil due to
the initially low pH might provide enough inorganic
2.6
2.4
2.2
pH
2.0
Set (A)
Set (B)
1.8
Set (C)
Set (D)
1.6
0 50 100 150 200 250 300
Time (hours)
Fig. 4 Variation in pH values in the leaching experiments with
A. ferrooxidans under various conditions over time. Set A,
bioleaching of A. ferrooxidans with inorganic nutrients and
Fe2+; set B, bioleaching of A. ferrooxidans with Fe2+; set C,
bioleaching of A. ferrooxidans with inorganic nutrients; set D,
bioleaching of A. ferrooxidans with water
Environ Geochem Health

nutrients to support the normal metabolism of A. where no inorganic nutrients were initially provided,
ferrooxidans. This result is of economic significance, there was a higher production of dissolved Fe3+ than
as the expense for inorganic nutrients externally in set A. Approximately 2.4 g/l of dissolved Fe3+ was
supplied to the bacterium can be reduced. additionally present in set B after 150 h compared
On the contrary, in experimental sets C and D, with set A. No initial supply of inorganic nutrients
little variation in the pH was observed over time once resulted in the inhibition of Fe precipitates, and, thus,
the pH had leveled out after the beginning of the a higher amount of dissolved Fe3+ in solution.
experiment. Ferrous ions, the essential energy source The amounts of Co extracted in sets A and B were
for A. ferrooxidans, was not supplied to either set; slightly greater than those in sets C and D, where no
therefore, unlike inorganic nutrients, a sufficient Fe2+ was supplied, although the difference dimin-
amount of Fe2+ to support bacterial metabolism ished after 250 h (Fig. 6). In comparison, the Sr
seemed to be naturally depleted in the studied soil. concentrations showed no significant difference
The iron speciation over time, i.e., dissolved total between the four batch-type slurries studied. The
Fe and Fe2+, in experimental sets A and B is depicted amounts of Sr extracted were almost the same in all
in Fig. 5. The amounts of Fe that formed precipitates sets, and appeared to be unaffected by the various
and colloidal suspensions in sets A and B were experimental conditions.
calculated to be 5.3 and 2.9 g/l respectively, after The extractions of Co and Sr were also signif-
150 h. According to Eq. 5, which indicates that Fe icantly influenced by the initial pH. The Co and Sr in
precipitation leads to a pH decrease, the result the artificially contaminated soil might be adsorbed
corresponded with the difference in solution pHs onto the surface of soil particles, and more easily
shown in Fig. 4. Interestingly, in experimental set B, dissolved by the low initial pH. Furthermore, the

10
30
Dissolved Co concentrations (mg/L)

Set (A) Total Fe

Dissolved Fe concentrations (g/L)

8 Fe2+ 25

20
6 Fe in precipitates and
colloidal suspensions 15
4 Set (A)
10
Set (B)
2 5 Set (C)
3+
Dissolved Fe Set (D)
0
0 0 50 100 150 200 250 300
0 50 100 150 200 250 300
Time (hours)
Time (hours)
30
Dissolved Sr concentrations (mg/L)

10
Set (B) Total Fe
25
Dissolved Fe concentrations (g/L)

Fe2+
8
Fe in precipitates and 20
colloidal suspensions
6 15

Set (A)
10
4 Set (B)
3+ 5 Set (C)
Dissolved Fe
2 Set (D)
0
0 50 100 150 200 250 300
0
0 50 100 150 200 250 300
Time (hours)
Time (hours)
Fig. 6 Variations in the dissolved Co and Sr concentrations in
Fig. 5 Variations in the aqueous total Fe and Fe2+ concentra- the leaching experiments with A. ferrooxidans under various
tions for bioleaching under various conditions over time conditions over time

123

formation of Fe precipitates and colloidal suspensions
probably acted as strong adsorbents, and, thus, there
were no increases in the amounts of dissolved Co and
Sr in the slurries over time.
The amounts of Fe colloidal suspensions in the
final solution after the 300 h of the experiment were
approximately calculated for the four slurries, accord-
ing to Fe mass balance equation, as follows; Fe in
colloidal suspensions = (Fe in the original soil + Fe in
the bacterial inoculum + Fe in the solution B of 9 K
medium)  (Fe in the residual soil after bioleach-
ing + dissolved Fe in the final solution). When Fe2+
was supplied to A. ferrooxidans, the amounts of Fe
colloidal suspensions were higher than those of Fe
precipitates (Fig. 7). In the practical application of a
bioleaching technique for the removal of radionuc-
lides from contaminated soil (e.g., soil washing), the
colloidal suspensions probably adsorb the released
radionuclides, which can be easily separated from the
residual soil, and then treated similarly to dissolved
radionuclides.
More Co than Sr was extracted. The difference
between the two elements was also reflected in the
analytical results obtained for the residual soils after
the bioleaching process. The residual soils were
digested with aqua regia, and the Co and Sr
concentrations determined. Under a range of given
experimental conditions, Sr remained in the residual
soils at higher concentrations than Co (Fig. 8).
It is remarkable, according to the final percentages
of Co and Sr removals from the contaminated soil,
that experimental sets A and B, with Fe2+ added,
showed almost equal or lower removals than sets C
and D. Because sets C and D did not contain large
amounts of initially supplied Fe2+, no significant Fe
5000
Fe in colloid
Fe in residual soil
4000
Fe mass (mg)
3000
2000
1000
0 radionuclide pollution through fall-out. Furthermore,
A B C D
Experimental sets
Fig. 7 Iron mass in colloidal suspensions and residual soil;
calculated using a mass balance equation
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Environ Geochem Health
100
88
Co Sr 90 91
Percentage of removal (%)
77
80
72
65
60
40
32 36
20
0
A B C D
Experimental sets
Fig. 8 The percentage Co and Sr removals under various
experimental conditions. The numbers near each bar represent
the percentage removal
precipitates or colloidal suspensions were formed;
therefore, no re-adsorption of extracted Co and Sr
onto solid Fe compounds could take place. In
particular, Sr showed a much lower extent of removal
when Fe2+ was externally supplied. While A. ferro-
oxidans supplied with Fe2+ extracted only 32–36% of
the Sr, the bacterium with no Fe2+ initially supplied
removed 65–77% of the Sr from the soil. Strontium
may be adsorbed more by Fe solids or precipitated
after the formation of compounds associated with
dissolved anions, such as sulfate.
This result suggested that preliminary investiga-
tions are required on the type of contaminant
radionuclides and the chemical and mineralogical
composition of the contaminated soil to be leached
for a cost-effective treatment to be implemented.
Since the reduced form of Fe (Fe2+) is essential for
the normal metabolism of A. ferrooxidans, sufficient
Fe2+ should be externally provided to the bacterium
to achieve satisfactory results from microbial metal
extraction. To date, a number of results, with high
efficiency of microbial metal lixiviation, have been
documented for low-graded sulfide ore minerals, and
even for soil and sediment contaminated with heavy
metals. However, according to the results presented
in this study, such good leaching efficiency may not
be expected when the contaminants are present in an
adsorbed mode in the soil or sediment substrate; e.g.,
in such cases, the initial supply of Fe2+ to stimulate
the Fe-oxidizer will form Fe solids, which re-adsorb
the metals leached into the solution. Only low pH
conditions may be adequate to efficiently extract such
Environ Geochem Health
weakly bound metals. Further investigations will be
required to examine the metal leaching efficiency of
leaching agents with a low pH and low Fe3+
concentrations, which can be easily achieved by the
simple addition of a weak acid.
Conclusion
Bioleaching and abiotic leaching of soil contaminated
with Co and Sr were conducted. In all cases, the
removal of Co and Sr from the soil was facilitated at
an initially low pH, as Co and Sr were present in a
loosely bound mode, via adsorption, onto the soil
surface. The extent of Sr removal was lower than that
of Co. A. ferrooxidans showed no significant
enhancement for the extractions of Co and Sr
compared with the abiotic control, although the
bacterium produced many protons and much dis-
solved Fe3+. Due to re-adsorption onto the Fe
precipitates and colloidal suspensions readily formed
by A. ferrooxidans in the case of Fe2+ addition, the
amounts of dissolved metals were almost equal to or
lower than those when no Fe2+ was supplied.
The conventional bioleaching technique, using Fe
oxidizers, e.g., A. ferrooxidans, may show no enhanced
efficiency for removal of radionuclides from natural
soils when supplied with sufficient Fe2+. This result
indicated that preliminary investigations are required
on the chemical and mineralogical compositions of the
contaminated soil to be leached, as well as the mode of
occurrence of radionuclides or heavy metals, for a
cost-effective treatment to be implemented.
Acknowledgement We appreciate the critical comments of
an anonymous reviewer, which greatly improved this
manuscript. This work has been financially supported by a
grant from the KESRI (Korea Electrical Engineering and
Science Research Institute; R-2003-B-351) to J.-U. Lee, which
was funded by the MOCIE (Ministry of Commerce, Industry
and Energy), Korea.
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