Biotechnology Letters 21: 715–718, 1999.

© 1999 Kluwer Academic Publishers. Printed in the Netherlands.

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Attachment behaviour of Thiobacillus ferrooxidans cells to refractory gold

concentrate particles

R. González, J. C. Gentina∗ & F. Acevedo

School of Biochemical Engineering, Universidad Católica de Valparaíso, Casilla 4059, Valparaíso, Chile
∗ Author for correspondence (Fax: (56-32) 273803; E-mail: jgentina@ucv.cl)

Received 30 June 1999; Accepted 2 July 1999

Key words: bioleaching, biooxidation, enargite, gold recovery, pyrite

Abstract
In the biooxidation of minerals, cells of Thiobacillus are distributed between the surface of the particles and the
liquid. This work quantifies the kinetics of attachment, the equilibrium between attached and suspended cells, and
the influence of ferric ion and particle size on this phenomenon. The attachment kinetics were fast, the equilibrium
was reached in 10 to 30 minutes, depending on the initial population. The equilibrium curves showed three distinct
phases, and the first two could be modelled by Langmuir equations. The maximum concentration of attached cells
increases with the addition of ferric ions and decreases with particle size.

Introduction ues. They suggested that the adsorbed ferrous ions on

Biohydrometallurgy has become a very attractive
technology to pretreat refractory gold concentrates.
Thiobacillus ferrooxidans, one of the most important
bioleaching microorganisms, bases its action on two
mechanisms, namely the direct and the indirect mech-
anisms (Torma 1977). The first one refers to sulphur
and sulphide oxidation by the bacteria attached to the
particle surface. The latter is related to the oxidation
of sulphur compounds by ferric ions resulting from
the bacterial oxidation of ferrous ions. Although it
is not easy to define the predominance of one or an-
other mechanism, the importance of cell attachment to
the particles in order to carry out the biooxidation is
widely accepted.
Cell attachment may be explained as the result of
interactions between microorganisms and active sites
in the particle surface, caused by electrostatic forces
or by the high internal energy of sites with large crys-
tallographic dislocations (Bagdigian & Myerson 1986,
Blake et al. 1995, Karavaiko et al. 1995).
Karavaiko et al. (1995) showed that the elec-
trophoretic motility of the Thiobacillus ferrooxidans
depends upon its physiological state and type of en-
ergy source, having a negative charge at low pH val-
the cell surface provide the charge difference between
cell and mineral particles.
Blake et al. (1995) analysed the Z potential of
Thiobacillus ferrooxidans, of different types of min-
eral particles and the relation between them. In this
case the cells also showed a negative Z potential,
which is reversed as a consequence of ferric ion ad-
sorption. Minerals have a negative net charge, while
the cell-particle complex tends to be neutral.
Furthermore, Ohmura et al. (1993) found a neg-
ative influence of ferrous and ferric ions on cell at-
tachment to different mineral particles. It was also
shown that cell attachment to sulphur minerals is se-
lective and that the hydrophilic interactions are of low
relevance.
Both, mineral particles and cells, are charged and
are able to adsorb ions from the solution. The charac-
teristics and concentrations of the ions will affect the
interactions between cells and particles.
The aim of this work is to characterise the at-
tachment of Thiobacillus ferrooxidans cells onto the
particles surface of a refractory gold concentrate and
to determine its dependency on ferric ion concentra-
tion, particle size and suspended cell concentration in
the liquid phase.
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Fig. 1. Kinetics of cell attachment on refractory gold concentrate,
particles size between 35–75 µm, at pH 1.5 and 33 ◦ C, starting with
different total amount of cells.
Experimental
Microorganism and culture conditions
Strain R-2 of Thiobacillus ferrooxidans, kindly sup-
plied by the Laboratory of Microbiology, Universidad
Católica de Chile, Santiago, was used in all exper-
iments. The microorganism was cultivated in a 9 K
medium (Silverman & Lundgren 1959), at pH 1.5 and
33 ◦ C, in a 3-litre aerated and mechanically agitated
bioreactor.
Mineral
The refractory gold concentrate was supplied by Min-
era El Indio (La Serena, Chile). Its main constituents
were (w/w): chalcopyrite (3.9%), chalcocite (0.8%),
covellite (0.3%), enargite, Cu3 AsS4 , (41%) and pyrite
(43%). Gold content was 42 g per tonne. Experiments
were run with three different particle size fractions:
35–53, 53–75 and 75–106 µm.
Experimental procedure
The attachment experiments were run in 100 ml Er-
lenmeyer flasks containing 15 ml of ferrous ion-free
9 K medium and with different concentrations of fer-
ric ion and cells. The pH was 1.5 and the temperature
was 33 ◦ C. The propagated cells were filtrated, cen-
trifuged, washed and resuspended to different concen-
trations. Total microbial protein was measured before
adding 0.93 g of mineral particles of each size frac-
tion. The flasks were placed in an agitated isothermal
rotary shaker for 3 hours to reach the equilibrium be-
tween attached an non-attached cells. Next, a liquid
sample was taken and centrifuged at 1000 g to elimi-
nate suspended mineral particles and it was analysed
for non-attached microbial protein. Attached micro-
bial protein was quantified as the difference between
total and non-attached cell protein. At the experimen-
tal conditions it was observed that cell growth and cell
attachment to the glass surface was negligible.
Analysis
Microbial protein was measured by alkaline hydroly-
sis of the cell envelopes and the Lowry method.
Results and discussion
Kinetics of cell attachment
Thiobacillus ferrooxidans cells attach to the surface of
mineral particles, reaching very promptly an equilib-
rium condition between suspended cells in the liquid
phase and cells attached to the particles. A typical ki-
netics of cell attachment is shown in Figure 1. Because
cell attachment is a fast phenomenon, it is not likely
that it will interfere with the overall process rate. How-
ever, it becomes important to study the conditions that
affect the equilibrium and its characteristics.
Influence of particle size and ferric ion concentration
on cell attachment
In order to analyse the influence of these parameters on
the attaching phenomenon, equilibrium curves were
generated relating attached cell concentration (XS )
and cell concentration in the liquid phase (XL ) at equi-
librium at defined experimental conditions. Figure 2 is
an example of the type of curves obtained in this kind
of experiment with and without ferric ion at different
particle sizes. In all the cases the curves have similar
shape, being possible to identify three zones in the
relationship between XS and XL .
Zones I and II resemble a two-step attachment phe-
nomena. Zone III is characterised by a decrease in XS
in response to higher values of XL . Neither the pres-
ence of ferric ion nor the changes in particle size affect
the shape of the curves.
The equilibrium data are strongly affected by ferric
ion. A lot more cells are observed attached to the solid
particles when 2 g l−1 of ferric ion are added to the
system. The positive charge of ferric ions interacting