Hydrometallurgy 59 2001.

177185
www.elsevier.nlrlocaterhydromet

Direct versus indirect bioleaching

Helmut Tributsch )
Dept. Solare Energetik, Hahn-Meitner Institut, 14109 Berlin, Germany
Received 19 October 1999; accepted 7 March 2000

Abstract
The dissolution of metal sulfides is controlled by their solubility product and thus, the wHqx concentration of the solution,
and further enhanced by several chemical mechanisms which lead to a disruption of sulfide chemical bonds. They include
extraction of electrons and bond breaking by wFe 3q x, extraction of sulfur by polysulfide and iron complexes forming
reactants wYqx and electrochemical dissolution by polarization of the sulfide whigh Fe 3q concentrationx. All these
mechanisms have been exploited by sulfide and iron-oxidizing bacteria. Basically, the bacterial action is a catalytic one
during which wHqx, wFe 3q x and wYqx are breaking chemical bonds and are recycled by the bacterial metabolism. While the
cyclic bacterial oxidative action via wHqx and wFe 3q x can be called indirect, bacteria had difficulties harvesting chemical
energy from an abundant sulfide such as FeS 2 , the electron exchange properties of which are governed by coordination
chemical mechanisms extraction of electrons does not lead to a disruption of chemical bonds but to an increase of the
oxidation state of interfacial iron.. Here, bacteria have evolved alternative strategies which require an extracellular polymeric
layer for appropriately conditioned contact with the sulfide. Thiobacillus ferrooxidans cycles wYqx across such a layer to
disrupt FeS 2 and Leptospirillum ferrooxidans accumulates wFe 3q x in it to depolarize FeS 2 to a potential where electrochemical oxidation to sulfate occurs. Corrosion pits and high resolution electron microscopy leave no doubt that these mechanisms
are strictly localized and depend on specific conditions which bacteria create. Nevertheless, they cannot be called direct
because the definition would require an enzymatic interaction between the bacterial membrane and the cell. Therefore, the
term contact leaching is proposed for this situation. In practice, multiple patterns of bacterial leaching coexist, including
indirect leaching, contact leaching and a recently discovered cooperative symbiotic. leaching where contact leaching
bacteria are feeding so wastefully that soluble and particulate sulfide species are supplied to bacteria in the surrounding
electrolyte. q 2001 Elsevier Science B.V. All rights reserved.
Keywords: Metal sulfides; Bioleaching; Thiobacillus ferrooxidans; Leptospirillum ferrooxidans; Interfacial kinetics

1. Introduction
The mechanism of bacterial leaching of sulfides
has interested researchers for a long time w1,2x. While
many parameters affecting the leaching process have

Fax:q49-30-8062-2434.
E-mail address: tributsch@hmi.de H. Tributsch..

been known as empirical factors, a more profound

understanding of the heterogeneous mechanism of
sulfide degradation has only gradually emerged as a
consequence of advances in semiconductor electrochemistry, sulfide surface chemistry and microbiological understanding. In order to face the question
whether bacteria are dissolving sulfides by direct
contact with their cell or by just recycling chemical
agents which disrupt the sulfide interface, the follow-

0304-386Xr01r$ - see front matter q 2001 Elsevier Science B.V. All rights reserved.
PII: S 0 3 0 4 - 3 8 6 X 0 0 . 0 0 1 8 1 - X

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H. Tributschr Hydrometallurgy 59 (2001) 177185

ing strategy should be assumed. First the possible

mechanisms of sulfide dissolution, as recognized by
modern sulfide electrochemistry should be reevaluated. Then, the possible strategies for bacteria aiming
at harvesting chemical energy should be discussed
and confronted with what is experimentally known
from studies on bacteriarsulfide interaction.

2. (Electro)chemical mechanisms of sulfide dissolution

Most sulfides are semiconductors or semi-metals,
the interfacial reactivity of which with electrolyte
species can be described in terms of electrochemical
potentials, electronic states, energy positions, energy
bands and electron exchange properties. From such
semiconductorelectrochemical studies of sulfides, it
is well known that depending on the limiting factors
involved, the electrochemical behavior and the nature of anodic corrosion products can be quite different. Many sulfides e.g. ZnS, CdS, PbS, CuS. yield
molecular sulfur as an oxidation product, others yield
sulfate e.g. FeS 2 , MoS 2 , WS 2 .. Still, another sulfide RuS 2 . does not corrode but remains kinetically
stable while evolving oxygen from water. Today, it
is quite well known why these sulfides are reacting
so differently, a critical factor being the electronic

nature of the valence energy band the highest energy band occupied by electrons. with which the
electrolyte may exchange electrons during the dissolution reaction Fig. 1..
While an extracted electron means a disrupted
bond when the valence band is derived from sulfur
S 2y states Fig. 1, left., no bond relevant for stability is broken when an electron is extracted from a
valence band derived from metal d-states Fig. 1,
right.. In this latter case, when metal centered electron transfer occurs the sulfur states are situated
approximately 1-eV lower and are not involved in
the reaction., an extracted electron means an increased oxidation state of interfacial metal centers.
This is the origin of a coordination chemical behavior of the sulfide interface.
The metal center reacts with water species from
the electrolyte or other small molecules forming
higher oxidized metal complexes Fig. 2..
Such a behavior, the attachment of water species
to the iron center, could clearly be simulated in
model experiments using low energy ion scattering
techniques w3x. It is also in agreement with the
observation that Ru in RuS 2 can oxidize water to
oxygen without corroding. With FeS 2 , however,
which has exactly the same crystal structure and the
same electronic structure, sulfate is formed when the
oxidative process is enhanced, since Fe cannot reach

Fig. 1. Energy scheme and examples of sulfides, which allow bond breaking in valence band or surface states. by electron extraction, left.
and of sulfides in which electron extraction only increases the oxidation state of the metal electron extraction from non-bonding metal
d-states..

H. Tributschr Hydrometallurgy 59 (2001) 177185

179

Fig. 2. Energy scheme of pyrite in contact with an electrolyte, showing how the energetic position of energy bands shifts with increasing
electrochemical potential due to complex formation of surface iron with water species, before a surface reaction takes place which involves
S 2y to yield thiosulfate, and finally sulfate.

higher oxidation states. This clearly shows that sulfate formation via a thiosulfate intermediate is the
consequence of a secondary reaction of hydrated and
oxidized surface iron with pyrite sulfur and not the
direct consequence of a reaction of water with pyrite
sulfur.
There are several mechanisms which can lead to a
disintegration of metal sulfides, converting them from
a crystalline state to soluble or amorphous products
Fig. 3.: protons can react with a sulfide Fig. 3a.
and liberate SHy ions, a reactivity defined by the
solubility product K so s wS 2y xwMe 2q x. When electrons are extracted by Fe 3q from a sulfide with S 2y
valence band Fig. 1, left side., chemical bonds are
broken and metal ions as well as molecular sulfur are
liberated Fig. 3b.. Broken chemical bonds can already be present in the sulfide p-type conduction,
low free energy of electrons.. Their presence in the
sulfide leads to a higher rate of interfacial dissolution
Fig. 3c.. Another way to break up a sulfide would
be to allow either a polysulfide or a metal complex
forming agent to interact with it Fig. 3d.. The
energy liberated during complex formation may be
sufficient to disintegrate the crystalline compound.
A further mechanism which can dissolve a sulfide
is a purely electrochemical one. When the electrical

potential of a sulfide is increased to the decomposition potential, it will react with the electrolyte.
In the case of pyrite, which is subject to a coordination chemical interfacial mechanism, water species
will first react with surface iron to form a complex
with stepwise increasing oxidation state. Since in
contrast to interfacial Ru in RuS 2 . Fe cannot form a
peroxo complex to liberate oxygen from water, it
undergoes a surface reaction with sulfur which finally leads to iron sulfate Fig. 3e..

3. Bacterial adaptations to chemical energy harvesting

Bacteria which evolved for harvesting inorganic
chemical energy from sulfides have only the choice
of relying on Fe 2q or on sulfur species for energy
supply. The faster the sulfide corrosion proceeds, the
more energy per time will be available, provided the
products are rich in energy. This is the case for HSy,
S o , S 2 O 32y, and Fe 2q; however, not for SO42y or
Fe 3q. In order to understand how bacteria have
adapted to the utilization of sulfide energy, it is now
helpful to understand on what most relevant factor
bacterial leaching with different sulfides depends, in

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H. Tributschr Hydrometallurgy 59 (2001) 177185

Fig. 3. Bond breaking mechanisms during sulfide degradation. Five different mechanisms are shown which lead to disintegration of sulfides.

presence of an iron-free nutrient solution. Comparative experiments have been performed with 16 different synthetic metal sulfides w4,5x. It turned out that
bacterial activity of Thiobacillus ferrooxidans and
thus, the rate of bacterial dissolution is approximately proportional to the solubility product of the
sulfide concerned Fig. 4..
This result is reasonable. This solubility product
of sulfides, K so , which spans 37 orders of magnitude, describes the reactivity of protons in disintegrating the sulfide into SHy and metal ions. SHy, of
course is a product which bacteria can oxidize for
energy recovery. This study has however also re-

vealed that additional mechanisms can increase sulfide dissolution. They involve situations or processes
in which bonding electrons are missing p-type conducting sulfidesFig. 3c., or where electrons are
extracted by Fe 3q Fig. 3b. so that the interfacial
bonds of the sulfide crystals are disrupted. In Fig. 4,
arrows are schematically indicating how these additional mechanism summarized in Fig. 3. are supporting bacterial activity. Fe 3q ions, which are extracting electron from sulfides supply bacteria with
chemical energy from Fe 2q and in addition, dissolve
the sulfide. However, as explained in Fig. 1, this is
only the case when bonding electrons are extracted

H. Tributschr Hydrometallurgy 59 (2001) 177185

181

Fig. 4. Bacterial leaching activity of T. ferrooxidans and mechanisms which improve leaching by additional bond breaking, in dependence
of the solubility product of the sulfide involved.

either from the sulfur-valence band of sulfides or

from surface states, which have shifted up from a
sulfur energy band into the forbidden energy region
as a consequence of proton reactions such a process
is, for example, relevant for the dissolution of ZnS
which has a large forbidden energy gap and a low
positioned sulfur energy band..
The interfacial reactions of FeS 2 and other sulfides listed in Fig. 1, right. are determined by the
coordination chemistry of the metal, and electron
extraction does not directly lead to the disintegration
of the sulfide. This explains why FeS 2 is quite
stable. Bacteria had to evolve specialized mechanisms to disrupt this abundant sulfide in order to get
access to its chemical energy.

T. ferrooxidans acquired the ability to use a polysulfide forming intermediate mechanism in Fig. 3d.
which is able to disrupt the pyrite. Recently, it has
been found that this carrier molecule works with a
thiol-group SHy. provided by the amino acid cysteine w6x. Control experiments have in fact shown
that cysteine alone can dissolve pyrite.
Leptospirillum ferrooxidans, which can only oxidize Fe 2q, however, not sulfur or SHy, apparently
learned to dissolve pyrite electrochemically. Electron
extraction itself does not break chemical bonds in
pyrite. However, when many electrons are extracted,
the electrical potential of pyrite moves so positive
that electrochemical formation of thiosulfate and sulfate occurs. This means the highly oxidized interfa-

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H. Tributschr Hydrometallurgy 59 (2001) 177185

under conditions where T. ferrooxidans already has

energy problems.
Summarizing the formal catalytic chemical mechanisms which bacteria are using for sulfide oxidation, we can in agreement with Ref. w5x, write:
MeS q 2 X q 2O 2
bacteria, chemicals

Me 2qq 2 X q SO42y

cial iron complex gets involved in a surface reaction

converting interfacial S 2y into sulfate Figs. 2 and
3e..
L. ferrooxidans has apparently learned to increase
the oxidation potential by applying a sufficiently
large concentration of Fe 3q to the sulfide according
to the well-known Nernst formula E s E o y
RTrzF .lnFe 3qrFe 2q .. It thereby uses electron extraction for depolarization and the electrochemical
dissolution of pyrite. Since it does not harvest sulfur
species and since it is able to gain redox energy from
Fe 2q at a quite positive redox potential, it can live

1.

With X s Hq acid leaching., X s Fe 3q ferric

iron leaching. and 2 X s Yqq Hq polysulfide car-

Fig. 5. Contact leaching, during which bacteria actively condition the FeS 2 interface by providing an extracellular polymeric layer EPL. as
reaction medium: a. T. ferrooxidans during extraction of sulfur, b. L. ferrooxidans during depolarization of FeS 2 using high Fe 3q
concentrations.. TEM pictures are shown to visualize the respective EPL which contain sulfur colloids in case a. and pyrite fragments in
case b. r10r.

H. Tributschr Hydrometallurgy 59 (2001) 177185

rier wYSHx mechanism., whereby the catalytic reactions can proceed simultaneously with different carriers or bacteria.
It is realized that the catalytic mechanisms involving Hq and Fe 3q can be considered to be indirect,
since the bacterial cells need not be in contact with
the sulfide, but only recycle the catalytic agent.
However, the situation is different when the sulfur
carrier Yq is employed T. ferrooxidans., or when
electron extraction by Fe 3q is used to electrochemically dissolve pyrite L. ferrooxidans..
On basis of these considerations, bacterial leaching activity BL . will be proportional to the solubility product K so . of the sulfide multiplied by an
enhancement factor E ., which considers increase of

183

leaching rate due to bond breaking mechanisms of

Fig. 3be, with E s E b Ec Ed Ee ; then:
B L ; K so E
2.
4. Contact leaching as redefined CdirectD leaching
In two discussed cases Fig. 3d,e., the bacteria
cannot rely on a chemical species as available in the
natural environment but have to produce an energy
carrier Yq. or have to increase artificially the concentration of an electron extracting agent Fe 3q .
close to the pyrite surface.
In both cases, bacteria had to create an artificially
controlled reaction zone as extracellular polymeric

Fig. 6. Corrosion pits formed by contact leaching of T. ferrooxidans in synthetic 100-nm-thin pyrite layer: a. from optical transmission
microscopy, b. from TEM picture.

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H. Tributschr Hydrometallurgy 59 (2001) 177185

layer between the bacterial membrane and the sulfide

Fig. 5..
In the case of T. ferrooxidans Fig. 5a., the
cysteine-based sulfur carrier is transported through it
to extract pyrite-sulfur to form a polysulfide, which
releases sulfur in colloidal form as a temporary
energy reservoir w7x. In the case of L. ferrooxidans,
Fe 3q is accumulated in the extracellular polymeric
layer to extract electrons at a sufficiently positive
potential to induce electrochemical thiosulfate and
sulfate formation. This mechanism may be compared
with the process of Aelectrochemical machining,B
where a metal block is contacted as anode and the
needle-shaped cathode is approached through the
electrolyte to produce localized anodic dissolution.
The localized corrosive bacterial action on a sulfide can easily be demonstrated. In the case of T.

ferrooxidans, the conditions can be selected to favor

localized activity which results in pronounced corrosion pits seen as bright dots perforating a synthetic
pyrite layer in optical transmission microscopic pictures. and chains of such pits, where bacteria have
multiplied w8x Fig. 6..
This means that the bacteria maintain a close
contact across the extracellular polymeric layer with
the sulfide interface in order to dissolve it.
Electron microscope pictures show the pits as
clearly locally etched nanocrystalline areas.
The original definition of direct leaching is a
leaching process by which the bacterial membrane
directly interacts with the sulfide using enzymatic
mechanisms. This is not the situation seen in electron
microscope pictures Figs. 5 and 6.. On the other
hand, even in a direct mechanism, it may be diffi-

Fig. 7. Scheme visualizing indirect leaching, contact leaching and cooperative leaching of a sulfide.

H. Tributschr Hydrometallurgy 59 (2001) 177185

cult to distinguish films of molecular dimensions

between bacterial cell wall and sulfide. For this
reason, it is suggested to rename the mechanism as
contact leaching. It would describe a situation in
which the bacterium attaches itself to the sulfide
with the purpose to condition the sulfide surroundings in such a way as to facilitate a dissolution
process which otherwise would not take place.

5. Bacterial leaching strategies

After having reached an overview over possible
sulfide leaching mechanisms, it remains to be discussed how bacteria have adapted to different sulfides and leaching environments. Some sulfides simply dissolve in acid solution or can be dissolved by
electron extraction with Fe 3q.
This is a favorable situation for indirect leaching
Fig. 1, left and Fig. 7., during which the bacteria do
not have to attach to the sulfide surface. Sulfides like
pyrite cannot easily be dissolved this way and contact leaching Fig. 5. has developed to break up the
crystal structure, either by sulfur extraction T. ferrooxidans. or by electrochemical dissolution L. ferrooxidans..
The higher the Fe 3q concentration in the leaching
solution becomes, the more Leptospirillum will be
favored, which only exists on electron extraction
paralleled by electrochemical dissolution of pyrite.
Simultaneously, with increasingly positive redox potential, T. ferrooxidans will gradually loose its ability to harvest chemical energy.
Recently, it has been shown that cooperative
leaching is possible w9x Fig. 7., during which bacteria attached to sulfide and free bacteria cooperate.
Thiobacilli ferrooxidans and thiooxidans. leach
pyrite and sulfur, respectively, while being attached
to the surface and thereby wastefully liberate energy
carrying species which feed bacteria which are in
suspension around. Such a behavior may have fa-

185

vored bacterial survival by providing maximum

chemical energy in presence of a limiting sulfide
surface.
6. Outlook
Bacteria have evolved multiple strategies for gaining chemical energy from sulfides of different electronic and crystalline structure. Besides indirect
leaching, contact leaching as well as cooperative
leaching with attached and suspended bacteria in
symbiotic activity is observed.
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