Documente Academic
Documente Profesional
Documente Cultură
HELMUT BRANDL
Zürich, Switzerland
1 Introduction 192
2 Terminology 192
3 Historical Background 192
4 Principles of Microbial Metal Leaching 194
4.1 Leaching Mechanisms 194
4.2 Models of Leaching Mechanisms 194
4.3 Factors Influencing Bioleaching 197
4.4 Bacterial Attachment on Mineral Surfaces 200
5 Microbial Diversity in Bioleaching Environments 200
6 Case Studies of Bioleaching Applications 208
6.1 Commercial-Scale Copper Ore Bioleaching 208
6.2 Reactor Bioleaching of Fly Ash 210
6.3 Shake Flask Bioleaching of Electronic Scrap 212
7 Economics of Metal Bioleaching 213
8 Perspectives of Bioleaching Technology 214
8.1 Heterotrophic Leaching 214
8.2 Leaching Under Thermophilic Conditions 215
8.3 Tapping Microbial Diversity 215
8.4 Treatment of Solid Wastes 215
8.5 Bioremediation of Metal-Contaminated Sites 215
9 Conclusion 217
10 References 217
192 8 Microbial Leaching of Metals
is also found in lead mines although it is of mi- mine in Spain 10 years earlier (SALKIELD,
nor value. In all these mines chrysocolla is also 1987). As a consequence to the ban of open air
artificially produced by slowly passing water ore roasting and its resulting atmospheric sul-
through the mine during the winter until the fur emissions in 1878 in Portugal, hydrometal-
month of June; subsequently, the water is evap- lurgical metal extraction has been taken into
orated in June and July. It is clearly demon- consideration in other countries more intense-
strated that chrysocolla is nothing but a de- ly. In addition to the ban, cost savings were an-
composed vein.” other incentive for the development: Heap
The German physician and mineralogist leaching techniques were assumed to reduce
Georgius Agricola (1494–1555) describes in transportation costs and to allow the employ-
his work de re metallica also techniques for the ment of locomotives and wagons for other ser-
recovery of copper that are based on the leach- vices (SALKIELD, 1987). From 1900 on, no open
ing of copper-containing ores (SCHIFFNER, air roasting of low-grade ore was conducted at
1977). A woodcut from his book illustrates the Rio Tinto mines.
the (manual) transport of metal-containing Efforts to establish bioleaching at the Rio
leachates from mines and their evaporation in Tinto mines had been undertaken in the begin-
the sunlight (Fig. 1). ning of the 1890s. Heaps (10 m in height) of
The Rio Tinto mines in south-western Spain low-grade ore (containing 0.75% Cu) were
are usually considered the cradle of biohydro- built and left for one to three years for “natu-
metallurgy. These mines have been exploited ral” decomposition (SALKIELD, 1987). 20 to
since pre-Roman times for their copper, gold, 25% of the copper left in the heaps were re-
and silver values. However, with respect to covered annually. It was calculated that ap-
commercial bioleaching operations on an in- proximately 200,000 t of rough ore could be
dustrial scale, biohydrometallurgical tech- treated in 1896. Although industrial leaching
niques had been introduced to the Tharsis operations were conducted at the Rio Tinto
mines for several decades, the contribution of
bacteria to metal solubilization was confirmed
only in 1961, when Thiobacillus ferrooxidans
was identified in the leachates.
Early reports state that factors affecting bio-
leaching operations were the height of the
heap, particle size, initial ore washing with
acid, and temperature control to about 50 °C
(SALKIELD, 1987). Another critical factor was
the supply of water for the leaching heaps. Al-
though usually acidic mine waters were used
for ore processing, 4 billion liters of freshwater
were required annually (SALKIELD, 1987).
Although metal leaching from mineral re-
sources has a very long historical record (EHR-
LICH, 1999; ROSSI, 1990) and although the oxi-
dation of reduced sulfur compounds and ele-
mental sulfur resulting in the formation of sul-
furic acid was demonstrated already in the
1880s (WINOGRADSKY, 1887), the oxidation of
metal sulfides was not described until 1922
when mobilization of zinc from zinc sulfide
Fig. 1. Woodcut from the book de re metallica writ- was investigated (RUDOLFS, 1922; RUDOLFS
ten by Georgius Agricola (1494–1555) illustrating and HELBRONNER, 1922). It was found that the
the manual recovery of copper-containing mine transformation of zinc sulfide to zinc sulfate
effluents which are collected in wooden basins and was microbially mediated. Based on these re-
concentrated in the sun. sults, the economic recovery of zinc from zinc-
194 8 Microbial Leaching of Metals
However, the model of “direct” and “indirect” FeS2c6 Fe3cc3 H2O ] S2O2c
3 P7 Fe
2c
a porin, and an iron(II)-sulfate chelate (BLAKE was autoclaved to obtain a sterile leaching so-
et al., 1993). The acid stability of rusticyanin lution without enzymatic activities and to eval-
suggests that it is located in the periplasmic uate the leaching ability of acid formed. (4)
space. Figure 2 shows a scheme of the model Leaching by fresh medium. Fresh non-inocul-
which combines the electron transport se- ated and sterile medium was added to the fly
quence proposed earlier with concepts stem- ash suspension and used as control. (5) Chem-
ming from the debate on “direct”/“indirect” ical leaching due to the preparation of the ash
leaching mechanisms (BLAKE and SHUTE, suspension (acidification to pH 5.4). Certain
1994; BLAKE et al., 1993; HAZRA et al., 1992; elements such as, e.g., Cd or Zn might be chem-
SAND et al., 1995). ically mobilized already during acidification.
Some details of the metal mobilization MWI fly ash contains reduced copper spe-
mechanism, the importance of the presence cies (chalcocite {Cu2S} or cuprite {Cu2O})
and attachment of microorganisms and their whereas zinc and others are present in their
active contribution have been demonstrated fully oxidized forms (BROMBACHER et al.,
for the leaching of fly ash from municipal 1998). Therefore, copper release from fly ash is
waste incineration (MWI) (BROMBACHER et directly affected and enhanced by T. ferrooxi-
al., 1998). Generally, several mechanisms of dans, whereas Zn, as well as Al, Cd, Cr, and Ni,
metal mobilization can be distinguished: (1) are released primarily due to the acidic envi-
Contact leaching effect on the release of met- ronment. Acidification of the fly ash pulp
als. Stock cultures of Thiobacillus ferrooxidans (chemical mobilization) led already to consid-
and Thiobacillus thiooxidans were added to erable extraction yields for Cd, Ni, and Zn and
ash suspensions and cells were in direct con- could slightly be increased using non-inoculat-
tact with the fly ash. Growth of thiobacilli ed sterile medium as lixiviant (Fig. 3). By com-
might be stimulated by increased energy avail- paring leached amounts of copper by filtered
ability from oxidation of reduced solid parti- cell free spent medium with autoclaved sterile
cles. (2) Metal solubilization by metabolically spent medium, it was concluded that signifi-
active (enzymatic) compounds in the absence cant amounts of copper were mobilized – in
of bacterial cells. Stock cultures were filtered contrast to other elements – by metabolic
to obtain the cell free spent medium. This me- products of T. ferrooxidans. Leaching with cell
dium was used for leaching. (3) Metal solubil- free spent medium indicating a solubilizing
ization by non-enzymatic extracellular meta- mechanism due to extracellular components
bolic products. Cell free spent medium (see 2) was significantly more effective than a leach-
ing with autoclaved spent medium where ex- ing (in g LP1): (NH4)2SO4 (3.0); K2HPO4 (0.5);
creted enzymes had been inactivated. It is MgSO4 · 7 H2O (0.5); KCl (1.0); Ca(NO3)2 ·
known that several components involved in 4 H2O (0.01); FeSO4 · 7 H2O (44.22); and 1 mL
the electron transport chain of Thiobacillus 10 N sulfuric acid (SILVERMAN and LUNDGREN,
(rusticyanin, cytochromes, iron–sulfur pro- 1959). Cells are harvested, diluted, and added
teins) are located in the periplasmic space to pyrite suspensions with a pulp density of
(BLAKE and SHUTE, 1994; SAND et al., 1995) 20 g LP1. Total soluble iron as well as sulfate
and might, therefore, also be present in the cell formed during oxidation is periodically deter-
free spent medium catalyzing oxidation of re- mined.
duced metal compounds. Metal bioleaching in acidic environments
In many leaching environments conditions is influenced by a series of different factors
(especially iron(II) and iron(III) concentra- (Tab. 1). Physicochemical as well as microbio-
tions) vary with the duration of the leaching. logical factors of the leaching environment are
This makes it difficult to assess the importance affecting rates and efficiencies. In addition,
and the effect of the presence of bacteria. Us- properties of the solids to be leached are of
ing an experimental setup to maintain con- major importance (ACEVEDO and GENTINA,
stant concentrations of ferrous and ferric iron, 1989; BRIERLEY, 1978; DAS et al., 1999; MURR,
it was possible to show that in the presence of 1980). As examples, pulp density, pH, and par-
T. ferrooxidans rates of pyrite or zinc sulfide ticle size were identified as major factors for
leaching are increased (HOLMES et al., 1999; pyrite bioleaching by Sulfolobus acidocalda-
FOWLER and CRUNDWELL, 1999; FOWLER et al., rius (LINDSTROM et al., 1993). Optimal condi-
1999). tions were 60 g LP1, 1.5, and ~20 µm, respec-
tively. The influence of different parameters
such as activities of the bacteria itself, source
4.3 Factors Influencing Bioleaching energy, mineralogical composition, pulp den-
sity, temperature, and particle size was studied
Standard test methods have been developed for the oxidation of sphalerite by T. ferrooxi-
to determine leaching rates of iron from dans (BALLESTER et al., 1989). Best zinc dis-
pyrite mediated by Thiobacillus ferrooxidans solution was obtained at low pulp densities
(ASTM, 1991). An active culture of T. ferro- (50 g LP1), small particle sizes, and tempera-
oxidans is grown in a defined medium contain- tures of approximately 35 °C.
198 8 Microbial Leaching of Metals
Tab. 1. Factors and Parameters Influencing Bacterial Mineral Oxidation and Metal Mobilization
Factor Parameter
manganese leaching efficiencies. At low con- It has been demonstrated recently that the
centrations of ~5 mg LP1, however, mangan- addition of small amounts of amino acids (cys-
ese mobilization was increased by 20% (BOUS- teine in this case) resulted in an increased
SIOS and MADGWICK, 1994). pyrite corrosion by T. ferrooxidans as com-
Also gaseous compounds can show inhibito- pared to controls without additions (ROJAS-
ry effects on metal leaching: Aqueous-phase CHAPANA and TRIBUTSCH, 2000). It is suggest-
carbon dioxide at concentration `10 mg LP1 ed that the microorganisms may profit from
was inhibiting growth of T. ferrooxidans on weakening and break up of chemical bonds
pyrite–arsenopyrite–pyrrothite ore (NAGPAL mediated by the formation of the cysteine–py-
et al., 1993). Optimal concentrations of carbon rite complex. This might also be the case under
dioxide were found to be in the range of 3 to natural conditions by the excretion of cys-
7 mg LP1. There are reports on the stimulation teine-containing metabolites. An inexpensive
of bacterial leaching and the increase of leach- alternative to increase metal recovery from
ing rates by supplementing leaching fluids with ore heaps by the addition of sulfur-containing
carbon dioxide (ACEVEDO et al., 1998; BRIER- amino acids such as cysteine has been suggest-
LEY, 1978; TORMA et al., 1972). Concentrations ed (TRIBUTSCH and ROJAS-CHAPANA, 1999).
of 4% (v/v) carbon dioxide in the inlet gas of a Other metabolites excreted by Thiobacillus
fermenter showed maximum growth rates of might also enhance metal leaching efficiencies:
T. ferrooxidans, maximum iron(II), copper, Wetting agents such as mixtures of phospho-
and arsenic oxidation (ACEVEDO et al., 1998). lipids and neutral lipids are formed by Thioba-
Pulp densities of 20 g LP1 delayed the onset cillus thiooxidans (BEEBE and UMBREIT, 1971).
of bioleaching of pyrite derived from coal As a consequence, growth of T. thiooxidans on
(BALDI et al., 1992). Increasing pulp densities sulfur particles is supported by the excretion
from 30 to 100 g LP1 decreased rates of pyrite of metabolites acting as biosurfactants which
oxidation in Sulfolobus cultures (NGUBANE facilitate the oxidation of elemental sulfur. It
and BAECKER, 1990). For fungi such as Asper- was also hypothesized that Thiobacillus caldus
gillus niger, optimal pulp densities for maxi- is stimulating the growth of heterotrophic or-
mum metal leaching efficiencies were found to ganisms in leaching environments by the ex-
be in the range of 30 to 40 g LP1 (BOSSHARD cretion of organic compounds and is support-
et al., 1996). Quartz particles at pulp densities ing the solubilization of solid sulfur by the for-
of 80 g LP1 almost completely inhibited the mation of surface-active agents (DOPSON and
oxidation of covellite by T. ferrooxidans espe- LINDSTROM, 1999). Metal solubilization might
cially in the absence of iron(II) (CURUTCHET also be facilitated by microbial metabolites ex-
et al., 1990). creted by organisms other than Thiobacillus
During bioleaching processes, coprecipita- which are part of microbial consortia found in
tion of metals with mineral phases such as ja- bioleaching operations. Microbial surfactants,
rosites can reduce leaching efficiencies (HI- which show large differences in their chemical
ROYOSHI et al., 1999). In addition, the precipita- nature, are formed by a wide variety of micro-
tion of compounds present in the leachates on organisms. In the presence of biosurfactants
the minerals to be leached can make the solid which lead to changes in the surface tension,
material inaccessible for bacterial leaching. metal desorption from solids might be en-
Organic solvents such as flotation or solvent hanced resulting in an increased metal mobil-
extraction agents, which are added for the ity in porous media. It has been suggested that
downstream processing of leachates from bio- this metabolic potential can be practically used
leaching, might also lead to inhibition prob- in the bioremediation of metal-contaminated
lems (ACEVEDO and GENTINA, 1989). Isopro- soils (MILLER, 1995). However, there is some
pylxanthate and LIX 984 (used as flotation evidence that surface-active compounds as
agent and solvent extraction agent, respective- well as organic solvents are inhibitory to bio-
ly) prevented the oxidation of pyrite and chal- leaching reactions and prevent bacterial at-
copyrite by T. ferrooxidans (HUERTA et al., tachment (MURR, 1980). The external addition
1995). This fact is of special importance when of Tween reduced the oxidation of chalcopyr-
spent leaching liquors are recycled for a reuse. ite by T. ferrooxidans (TORMA et al., 1976). It
200 8 Microbial Leaching of Metals
bioleaching habitats (SAND, 1992; SCHRENK et extraction from mine tailings is more efficient
al., 1998). Thiobacilli are members of the divi- using thermophilic instead of mesophilic or-
sion of Proteobacteria close to the junction ganisms, extremely thermophilic microorgan-
between the and subdivision whereas isms show a higher sensitivity to copper and to
leptospirilli are placed in the Nitrospira divi- high pulp densities in agitated systems limit-
sion (RAWLINGS, 1999). Genetic studies re- ing, therefore, some practical applications
vealed that the role of T. ferrooxidans in leach- (DUARTE et al., 1993; NORRIS and OWEN,
ing operations has probably been overestimat- 1993).
ed. Excellent reviews on the genetics of Thio- Although environmental conditions in
bacilli and leptospirilli have been published leaching operations favor the growth and de-
recently (RAWLINGS, 1999; RAWLINGS and KU- velopment of mesophilic, moderately thermo-
SANO, 1994). philic, and extremely thermophilic microbial
Thiobacillus ferrooxidans belongs to the communities, metal leaching at low tempera-
group of chemolithotrophic organisms. The or- tures has also been observed. Copper and
ganism is rod-shaped (usually single or in nickel were leached from pyritic ore samples
pairs), non-spore forming, gram-negative, mo- in significant amounts at 4 °C (AHONEN and
tile, and single-pole flagellated (HORAN, 1999; TUOVINEN, 1992). However, leaching rates
KELLY and HARRISON, 1984; LEDUC and FER- were lower by a factor of 30 to 50 as compared
RONI, 1994; MURR, 1980). As carbon source, to experiments conducted at 37 °C. T. ferrooxi-
carbon dioxide is utilized. Ferrous iron is oxi- dans recovered from mine waters was able to
dized. Ammonium is used as nitrogen source. grow at 2 °C with a generation time of approx-
Although T. ferrooxidans has been character- imately 250 h suggesting a psychrotrophic na-
ized as being a strictly aerobic organism, it can ture of the organism (FERRONI et al., 1986).
also grow on elemental sulfur or metal sulfides Bacterial iron mobilization has also been ob-
under anoxic conditions using ferric iron as served at 0 °C in ore samples obtained from
electron acceptor (DONATI et al., 1997; PRONK Greenland (LANGDAHL and INGVORSEN, 1997).
et al., 1992). Solubilization rates at these low temperatures
The genus Thiobacillus represents a versa- were still approximately 25 to 30% of the max-
tile group of chemolithoautotrophic organ- imum values observed at 21 °C. All these find-
isms. Optimum pH values for growth vary ings may have a potential for practical applica-
between 2 and 8 (Fig. 4). It has been demon- tions in geographical areas where field opera-
strated that sulfur-oxidizing bacteria are ca- tions are subjected to low temperature regimes.
pable of reducing the pH of highly alkaline fly A series of heterotrophic microorganisms
ash suspensions amended with elemental sul- (bacteria, fungi) is also part of microbial bio-
fur from approximately 9 to 0.5 (KREBS et al., leaching communities (Tab. 2). This group of
1999) (Fig. 5). It is likely that thiobacilli con- organisms uses extracellular metabolites and
tribute to increasing acidification of leaching cell lysates from autotrophs as carbon source
ecosystems in a successive mode: In the initial resulting in the removal of an inhibitory excess
stages the growth of less acidophilic strains of carbon and stimulating, therefore, growth
(e.g., Thiobacillus thioparus) is stimulated and iron oxidation of thiobacilli (BUTLER and
whereas during prolonged leaching the pH de- KEMPTON, 1987; FOURNIER et al., 1998). In ad-
creases gradually supporting growth of more dition, several heterotrophs can also con-
acidophilic strains. This has already been ob- tribute to metal solubilization by the excretion
served in metal leaching from wastewater sew- of organic acids such as citrate, gluconate, oxa-
age sludge (BLAIS et al., 1993). late, or succinate.
A variety of thermophilic microorganisms
(especially Sulfolobus species) has been en-
riched and isolated from bioleaching environ-
ments (BRIERLEY, 1990; NEMATI et al., 2000;
NORRIS and OWEN, 1993). Temperature optima
for growth and metal leaching were in the
range between 65 and 85 °C. Although copper
202
Tab. 2. Microbial Diversity of Acidic Bioleaching Environments and Acidic Mine Drainage: Selection of Microorganisms known to Mediate Metal Bio-
leaching Reactions from Ores and Minerals or known to be Part of the Microbial Consortia Found in Bioleaching Habitats
203
Pseudomonas cepacia heterotrophic 5.4–6.0 22 BAGLIN et al. (1992)
204
Tab. 2. Continued
205
206
Tab. 2. Continued