Minerals Engineering 48 (2013) 2024

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Minerals Engineering
journal homepage: www.elsevier.com/locate/mineng

Evaluation of bioleaching factors on gold recovery from ore by

cyanide-producing bacteria
Doyun Shin a,, Jinki Jeong a, Sujeong Lee a, B.D. Pandey b, Jae-chun Lee a
a
Mineral Resources Research Division, Korea Institute of Geoscience and Mineral Resources (KIGAM), Gwahangno 124, Yuseong-gu, Daejeon 305-350, Republic of Korea
b
Metal Extraction and Forming Division, CSIR-National Metallurgical Laboratory, Jamshedpur 831 009, India

a r t i c l e i n f o a b s t r a c t

Article history: The present study was conducted to investigate the gold bioleaching factors from ore by cyanide produc-
Available online 25 April 2013 ing bacterium Chromobacterium violaceum. The optimal condition for cyanide production by C. violaceum
was pH 9 and 5 g/L of glycine in YP medium in 2-days of incubation. In shake ask culture, gold bioleach-
Keywords: ing from the ores by C. violaceum was determined with various experimental protocols such as particle
Cyanide producing bacteria size, pre-grown C. violaceum, pH, and biooxidative treatment. The three types of low grade ores viz., R,
Chromobacterium violaceum S, and H were used. The gold bioleaching efciencies were recorded as 0%, 50%, and 5% for ores R, S,
Bioleaching
and H, respectively, when C. violaceum culture was used without any pretreatment (protocol 1). In the
Gold
Low-grade ore
experimental protocol involving grinding and pre-grown C. violaceum, leaching efciencies increased
to 60%, 100%, and 40% for ore R, S, and H samples, respectively. Especially, the bioleaching efciency of
ore S enhanced to almost 100% with pre-grown C. violaceum (protocol 2) and that of ore R increased to
53% (i.e., 96% of cyanidable gold) with grinding (protocol 3) due to their mineralogical characteristics
of the ores. For refractory gold (i.e., ore R) grinding as pretreatment was needed, and for ore S (almost
all of the gold was cyanidable) cyanide production was activated by using pre-grown bacteria. Biooxida-
tion with Acidithiobacillus ferrooxidans and pH adjustment (i.e., 911) did not affect the bioleaching ef-
ciencies. The mineralogical cause of gold refractoriness was analyzed by automated SEM that showed
most of gold in the ore was entrapped in pyrite and silica. The results indicated that gold bioleaching
by C. violaceum from low grade ore can be enhanced by grinding and pre-grown microbe; use of appro-
priate experimental condition is important according to the mineralogical characteristics.
2013 Elsevier Ltd. All rights reserved.

1. Introduction organic waste can be used as a cheaper substrate to generate cya-

Cyanidation to recover gold from ore or secondary resources has
been in use almost universally based on formation of gold-cyanide
complex which shows very good water solubility and high chemical
stability (Chadwick and Sharpe, 1966). However, excessive use of
cyanide for the dissolution of gold is associated with environmental
risk, and thus biological methods for gold leaching are being inves-
tigated as environmental friendly methods (Olson, 1994). Since cya-
nide producing bacteria autonomously detoxify cyanide to b-
cyanoalanine (Brysk et al., 1969), an integrated biological cyanida-
tion process without cyanide discharge can be designed involving
cyanide production and degradation simultaneously. Biological
cyanidation also has advantage of producing biogenic cyanide with-
out having transported the chemical to the processing site.
Cyanide producing bacteria such as Chromobacterium violaceum,
Bacillus megaterium, Pseudomonas uorescens, and Pseudomonas
plecoglossicida (Knowles and Bunch, 1986) produce cyanide from
glycine by oxidative decarboxylation (Blumer and Haas, 2000). If
nide instead of glycine, it will be possible to reduce the expenses in
the biological cyanidation process. Various other resources such as
ores (Campbell et al., 2001), electronic scraps (Brandl et al., 2008),
and printed circuit boards of waste electronics (Brandl et al., 2001;
Faramarzi et al., 2004; Tran et al., 2011a, 2011b) have also been
studied to recover gold by C. violaceum. However, few or no studies
have dealt with experimental conditions which are practically
important for gold bioleaching from low grade ores.
In this work the feasibility of gold bioleaching by C. violaceum,
from low-grade ores containing gold was evaluated with various
experimental conditions such as particle size, incubation tech-
nique, pH, and biooxidative treatment. In addition, the mineralog-
ical studies of the ores were performed to understand the cause of
refractoriness in the samples towards gold leaching.
2. Materials and methods
2.1. Microorganisms and medium

Corresponding author. Tel.: +82 42 868 3616. C. violaceum DSM 30191T and Acidithiobacillus ferrooxidans DSM
E-mail address: doyun12@kigam.re.kr (D. Shin). 14882T were purchased from DSMZ (Germany). C. violaceum was

0892-6875/$ - see front matter 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.mineng.2013.03.019
 D. Shin et al. / Minerals Engineering 48 (2013) 2024 21

grown in YP medium (Tran et al., 2011b) at 30 C with various pH
(i.e., 6.7, 8, 9, 9.5, 10, 11, and 12) and glycine concentration (i.e.,
0.5, 1, 2, 5, 10, and 20 g/L). Bacterial density was determined by
monitoring optical density at 600 nm and the cyanide concentra-
tion was analyzed by ion-selective electrode for cyanide and titra-
tion against standard AgNO3 solution following the EPA Method
9014. A. ferrooxidans was maintained at 30 C in the 9 K medium
(Silverman and Lundgren, 1959) which was lter-sterilized and
pH was adjusted to 1.8 with 0.1 N H2SO4.
2.2. Ore characterization
The ores used in this study were designated as R, S, and H. The
gold and silver contents of the ores were measured by re assay
and the copper content by ICP-AES after aqua regia digestion.
The mineralogy of the ores was analyzed by XRD and SEMMLA
(Table 1a and b). The ores was <1700 lm and further ground and
then sieved below 180 lm. Total gold, silver, and copper available
for cyanidation was measured by adding 300 mg/L of sodium cya-
nide solution (pH 10.8) into each ores (<180 lm) at 100 g/L of pulp
density and shaking until the leached gold concentration did not
increase.
2.3. Shake ask culture
Preliminary experiments demonstrated that the optimal pulp
density was 20 g/L and the sampling point was the fth day of
incubation because the gold, silver, and copper dissolution in-
creased until 4 days and reached maximum in 5 days. The proto-
cols used to investigate the experimental conditions are given
below.
Protocol 1: 30 g of the ore (<1700 lm) was added into 2-L of
Culture bottle containing 1.5 L of YP medium including 5 g/L of gly-
cine. Then 15 mL of pre-grown C. violaceum (approximately 108
CFU/mL) in YP medium was inoculated. The asks were shaken
on a shaking incubator (220 rpm) at 30 C for ve days.
Protocol 2: C. violaceum was inoculated into YP medium and
incubated overnight. In the pre-grown culture, the ore
(<1700 lm) and glycine were added and the experiment was run
in the same condition as in protocol 1.
Protocol 3: The experiment was run under the same condition
as in protocol 1, except for the size of the ore (i.e., 180 lm).
Protocol 4: The experimental condition was the same as in pro-
tocol 2, except for the size of the ore (i.e., 180 lm).
Protocol 5: The experimental condition was the same as in pro-
tocol 4, except for the pH adjustment (911) before adding the ore
(<180 lm) because the most effective pH for gold cyanidation has
been reported to be pH 10.511 (Rees and Van Deventer, 1999).
Protocol 6: The ore (<180 lm) was pretreated with A. ferrooxi-
dans for 2 months and then, the slurry containing treated ore in
the culture was ltered with 0.22 lm pore lter, washed with dis-
tilled water and dried at 70 C for overnight. The subsequent pro-
cedure that was followed was the same as in protocol 4. The
solid and liquid was separated by centrifuge at 3500 rpm for

2.0
2.0
1.8
(a) pH 6.7 (a) glycine 0.5 g/L
pH 8 1.8 glycine 1 g/L
1.6 pH 9 glycine 2 g/L
1.6
Bacterial growth (OD600)

pH 9.5
Bacterial growth (OD600)

glycine 5 g/L
1.4 pH 10 glycine 10 g/L
1.4
pH 11 glycine 20 g/L
1.2 pH 12 1.2
1.0
1.0
0.8
0.8
0.6
0.6
0.4
0.4
0.2
0.2
0.0
0.0
0 1 2 3 4 5
0 1 2 3 4 5
Incubation time (day) Incubation time (day)

50 50
(b) pH 6.7 (b) glycine 0.5 g/L
pH 8 glycine 1 g/L
Cyanide concentration (mg/L)
Cyanide concentration (mg/L)

pH 9 glycine 2 g/L
40 40
pH 9.5 glycine 5 g/L
pH 10 glycine 10 g/L
pH 11 glycine 20 g/L
30 pH 12 30

20 20

10 10

0 0
0 1 2 3 4 5 0 1 2 3 4 5
Incubation time (day) Incubation time (day)

Fig. 1. (a) Bacterial density and (b) cyanide production of C. violaceum with various Fig. 2. (a) Bacterial density and (b) cyanide production of C. violaceum with various
pH (i.e., 6.7, 8, 9, 9.5, 10, 11, and 12) in YP medium containing glycine 5 g/L. concentrations of glycine (0.5, 1, 2, 5, 10, 20 g/L) in YP medium at pH 9.
22 D. Shin et al. / Minerals Engineering 48 (2013) 2024

Cyanidable metal Cyanidable metal

100 (a) Gold
100 (c) Gold

leached metal (%)

leached metal (%)

Silver Silver

Cyanidable and

Cyanidable and
80 Copper 80 Copper

60 60

40 40

20 20

0 0
R S H R S H
Ore type Ore type

Cyanidable metal Cyanidable metal

100 (b) Gold
100 (d) Gold
leached metal (%)

leached metal (%)

Silver Silver
Cyanidable and

Cyanidable and
80 Copper 80 Copper

60 60

40 40

20 20

0 0
R S H R S H
Ore type Ore type

Fig. 3. Gold, silver, and copper bioleaching efciencies (% of gold, copper, and silver leached by C. violaceum against total gold, copper, and silver in the ores) at 20 g/L of pulp
density and pH 9 (a) in protocol 1; (b) in protocol 2; (c) in protocol 3; (d) in protocol 4.

10, little or no bacterial growth was observed. The highest cyanide

Table 1
production (i.e., 34.56 mg/L) was observed at pH 9 with 5 g/L gly-
(a) Chemical and (b) mineralogical analysis of the ores used in this study.
cine even though the bacterial growth was higher at pH less than
R S H 9. As shown in Fig. 2b, the cyanide production was similar between
The total content of metals in the ore (mg/kg) 2 and 5 g/L of glycine at 2 days, however, at day 1 the cyanide pro-
Panel (a) duction was the highest at 5 g/L of glycine level. Comparing with
Au 1.149 1.343 21.201
the cyanide production in Fig. 1b, 5 g/L of glycine was deemed to
Ag 13.9 <2.0 176.6
Cu 60 120 136 be the most appropriate concentration in cyanide production.
Therefore, pH 9 and 5 g/L of glycine was used throughout this
The cyanidable metal in the ore (mg/kg)
Au 0.63 0.00 1.21 0.07 6.18 1.49 study. The higher amount of cyanide produced in early 12 days
Ag 6.14 0.26 0.53 0 18.95 5.95 might be continuously used during the entire leaching period along
Cu 18.32 1.30 2.39 0.78 39.68 1.56 with the consumption of reduced cyanide being generated in the
R S H subsequent period.
Mineralogical composition of the oresa
Panel (b)
Quartz (SiO2)b Gibbsite (Al(OH)3) Quartz (SiO2) 3.2. Shake ask culture
Sanidine (K(Si3AlO8) Hematite (Fe2O3) Muscovite
(KAl2Si3AlO10(OH)2) The bioleaching efciencies of gold from the ores by C. violace-
Pyrite (FeS2) Goethite Pyrite (FeS2) um were determined in shake ask culture with various factors and
(FeO(OH))
the results are shown in Fig. 3ad. The bioleaching efciencies
Muscovite Kaolinite Kaolinite
(KAl2Si3AlO10(OH)2) (Al2Si2O5(OH)4) (Al2Si2O5(OH)4) throughout this study were expressed against the content of the
Magnetite (Fe3O4) Rutile (TiO2) Wollastonite (CaSiO3) total metal chemically analyzed. The cyanidable metals in
Anatase (TiO2) Table 1a were also presented in Fig. 3ad to compare the leaching
Magnetite (Fe3O4)
efciencies. As can be seen the leaching efciency of gold varied
a
The minerals were drawn in quantitative order. with the nature of the ore. The bioleaching efciency had increas-
b
Minerals in bold are major components. ing tendency in protocol 24 compared to that of protocol 1. Espe-
cially, gold bioleaching was enhanced appreciably in ore R by
30 min, and the metal concentrations in the leaching solution and
grinding (from 0 to 53%, between protocols 1 and 3) and ore S by
the residue were analyzed by AAS and re assay, respectively. By
using pre-grown C. violaceum (from 50 to 100%, between protocols
this procedure, 99109% of total metal was recovered. The leaching
1 and 2), respectively, probably because of the characteristics of
efciencies were normalized based on the mass balance. All exper-
the ores. It may be attributed to the fact that the refractory gold
iments were performed in triplicates.
in ore R would be liberated by grinding and then leached, whereas
3. Results and discussions almost all of the gold in ore S was not refractory so leached by acti-
vating bacterial cyanide production. The pre-grown C. violaceum
3.1. Cyanide production by C. violaceum and grinding did not affect the leaching of ore R and S, respectively.
In the case of ore H, the bioleaching efciency increased from 5% to
The growth and cyanide formation of C. violaceum at varying pH 18% by grinding (protocol 2) and to 40% by using pre-grown C. vio-
and glycine concentration are presented in Figs. 1 and 2. Above pH laceum and grinding (protocol 4). In protocols 5 and 6 (i.e., pH
 D. Shin et al. / Minerals Engineering 48 (2013) 2024 23

Fig. 4. Scanning electron photomicrographs of gold (white spots in the red circles) in ores R and H. The detailed investigations are given in the text. (For interpretation of the
references to colour in this gure legend, the reader is referred to the web version of this article.)

adjustment and biooxidation), the gold bioleaching did not in-
crease (data not shown). It may be because of the presence of very
small content of pyrite in these ores and the gold particles might be
entrapped in other minerals. Therefore, the mineralogical analysis
was performed in the following section.
3.3. Mineralogical causes of gold refractoriness
Based on the mineral liberation analysis (MLA) with FE-SEM,
one gold particle in free metallic state (the brightest spots in red
circles) was found in ore R and the elemental composition was gold
and silver (Fig. 4a). In ore H, some gold was identied as shown in
Fig. 4bf. All of the gold found in the ores was the natural alloy of
gold and silver (i.e., electrum). In Fig. 4b, gold spot consisted of gold
and silver and the dark gray part surrounding gold was pyrite. It
was expected that gold was entrapped in pyrite mineral
(Table 1b). However, pyrite in Fig. 4b was surrounded by SiO2 or
Al, O, Si, and K minerals. Free gold particle could also be seen in
Fig. 4c and the gold spots in submicroscopic size were observed
in Fig. 4df. The rest part of the particles was quartz (SiO2) in
Fig. 4d and the minerals which consisted of Al, O, Si, and K in
Fig. 4e and f. Based on the above results, it may be concluded that
the gold bioleaching efciency in ore H was not enhanced by bio-
oxidation because pyrite was not dissolved by A. ferrooxidans due
to the presence of minerals like quartz and other phases around
it. Ultra ne milling should be used to liberate pyrite and gold par-
ticles from the ores and then biooxidation can be applied.
4. Conclusions
In summary, this study evaluated the bioleaching factors of gold
from low grade ore by C. violaceum. The optimal condition for cya-
nide production was pH 9 and 5 g/L of glycine in this study. In
24 D. Shin et al. / Minerals Engineering 48 (2013) 2024
shake ask culture the highest gold bioleaching by C. violaceum
was observed in the experimental condition using pre-grown C.
violaceum and smaller particle size (i.e., 180 lm) at 20% pulp den-
sity. The low concentration of cyanide is still a technical subject so
further study is underway to develop cyanide capture system to
produce highly concentrated cyanide and reduce the amount of
glycine because of the cost and toxicity of glycine. These ndings
clearly suggest that biological gold leaching by C. violaceum can
be used as an alternative of chemical cyanidation if appropriate
pretreatment (i.e., grinding) or experimental technique (i.e., pre-
grown bacteria) is applied.
Acknowledgment
This research was supported by the Basic research Project of the
Korea Institute of Geoscience and Mineral Resources (KIGAM)
funded by the Ministry of Knowledge Economy of Korea.
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