The effect of growth medium of Thiobacillus ferrooxidans on pyrite flotation

Pergamon

THE EFFECT

MineralsEngineering,Vol.9, No. 2, pp. 157-168,1996 Copyright© 1996Publishedby ElsevierScienceLtd Printedin GreatBritain.All rightsreserved Plh S0892-6875(96)00001-5 0892-6875/96 $15.00+0.00

OF GROWTH MEDIUM OF THIOBACILLUS ON PYRITE FLOTATION

FERROOXIDANS

M. MISRA, K. BUKKA and S. CHEN Department of Chemical and Metallurgical Engineering, University of Nevada, Reno, NV 89557, USA

(Received 2 August 1995; accepted 18 October 1995)

ABSTRACT

The bacterium, Thiobacillus ferrooxidans, has been used for the depression of pyrite during the flotation separation of coal particles from associated minerals. However, the utilization of T. ferrooxidans as a regulator in the differential flotation of sulfide minerals is yet to be established. The conventional reagent used for pyrite depression is cyanide which is toxic and in many cases nonselective. The objective of the present investigation is to determine if T. ferrooxidans can be used for the selective depression of pyrite. In view of this objective T. ferrooxidans was grown in a variety of culture media to render bacteria with different surface properties such as contact angle and zeta potential. The passivity of pyrite surface due to sulfur coating, which is formed by the oxidation of the mineral, was examined using AC Impedance Spectroscopy. The influence of bacterial surface properties on adhesion and subsequent hydrophilic/hydrophobic balance of the pyrite mineral surface with regard to flotation recovery was examined. The flotation response of pyrite was interpreted in terms of surface modifications induced by the adsorption of bacteria.

Keywords Sulphide ores; froth flotation; surface modification; bacteria

INTRODUCTION The bacterium, Thiobacillusferrooxidans, has traditionally been used for the biooxidation of refractory gold ores and bioleaching of selected sulfide minerals. In recent years a modest amount of work has been directed to the selective depression of pyrite (FeS2) from complex coal-pyrite matrix [ 1-4]. Initially, pyrite present in coal is oxidized by biotreatment so that hydrophobic coal can be separated from hydrophilic pyrite by a variety of flotation methods [5]. It has been postulated that superficial surface oxidation and formation of hydrophilic sites on pyrite is responsible for depression [6]. However, the utilization of such a processing strategy for coal-pyrite depression is yet to be commercialized. In complex sulfide mineral flotation systems, the most often used depressant for pyrite is potassium and/or sodium cyanide. Cyanide is known to be toxic and environmental regulations are stringent for final disposal of cyanide bearing effluents. Due to the environmental regulations and constrictions, there is a need to develop an alternative pyrite depressant. One of the potentially promising approaches involves the use of

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biological reagents such as Thiobacillus ferrooxidans. The interaction of T. jerrooxidans with sulfide minerals has been the subject of numerous investigations [7-101. The underlying phenomenon which determines the pyrite-bacterium interaction is the attachment of the microorganism to the mineral surface. This attachment or adhesion is influenced by the surface properties of the bacterium which in turn are dependent on culturing conditions and composition of the growth media. In this paper the effect of different culture media on selective attachment and subsequent depression of pyrite is discussed. Although preliminary in nature, the studies have shown that by adjusting the growth medium the adhesion of the organism to pyrite can be increased and selective separation can be achieved. Changes in the electrical properties of pyrite as a result of superficial oxidation, which leads to the formation of elemental sulfur on the surface, is investigated by AC Impedance Spectroscopy. The impact of elemental sulfur formation on the hydrophobic/hydrophilic balance of the mineral surface, both in the absence and in the presence of T. ferrooxidans, is also discussed.

M A T E R I A L S AND M E T H O D S Microorganism, medium, and conditions of cultivation The iron oxidizing bacterium, Thiobacillus ferrooxidans, was obtained from American Type Culture Collection. The nutrient composition of the three different culture media viz., 9K Standard Medium (SM), Phosphate Deficient Medium (PDM), and Pyrite Added Medium (PAM) used for the growth of bacteria is presented in Table 1. The cultivation of T. ferrooxidans was carried out in four 250 ml. Erlenmeyer flasks which were placed in the shaker bath at 300C for four days. Iron-free cell suspensions were used in all preparations. For this purpose 1 L of a 4-day old bacterial culture was first passed through Whatman No. 44 filter paper in order to remove the precipitates. The filtrate was then spun in a refrigerated centrifuge (JZ-21 Beckman, SS-34 rotor) at 15,000 rpm for 30 minutes. The residual pellet was suspended in a solution of sulfuric acid at pH 1.8 and allowed to stand for 2 hours in a refrigerator to permit settling of any precipitate. The supernatant containing the cells was again centrifuged as described earlier. Washing with sulfuric acid was repeated until the cell suspensions were free from iron. The cell pellet was finally suspended in 150 ml sulfuric acid (pH = 1.8) and stored in a refrigerator at 4°C. The concentration of T. ferrooxidans was determined by a cell counting technique by using a polarizing optical (Nikon) microscope. T A B L E 1 Nutritional composition of the three different culture media used in the growth of Thiobacillus ferrooxidans 9K Standard Medium 3.0 g/L (NH4)2SO 4 0.5 g/L MgSO 4- 7H20 0.1 g/L KC1 0.5 g/L K2HPO 4 142.2 g/L FeSO 4. 7H20

Phosphate Deficient Medium

Pyrite Added Medium

3.0 g/L (NH4)2SO 4 0.5 g/L MgSO4.7H20 0.1 g/L KC1 42.2 g/L FeSO 4. 7H20

3.0 g/L (NH4)2SO4 0.5 g/L MgSO 4. 7H20 50.0 g/L Pyrite

Minerals The sample of pyrite was obtained from Ward's National Science Establishment. The purity of pyrite was reported to be > 95%. Contact Angle Measurement For measuring contact angles the mineral sample was cut into a 2 x 2x 2 cm cube and one of the surfaces was wet ground under a stream of deionized distilled water on grit papers #320 to #600. The sample was then wet polished on a broad cloth mounted on a plate using 1.0 t~m and 0.5tzm abrasive alumina paper. After careful polishing, the specimen was cleaned with a jet of deionized distilled water to remove dirt and

Effect of growth medium on pyrite flotation

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alumina powder. Prior to using in the contact angle measurement the sample specimen was polished with a clean broad flannelette. Plastic surgical gloves were used at all times to handle the specimen in order to avoid contamination. Contact angle measurements were made using a Rame-Hart Contact Angle Goniometer (Model No. 100). Freshly prepared T. ferrooxidans suspensions were pH adjusted with dilute sulfuric acid (pH =2.4) and conditioned together with pyrite sample on a rotary shaker at 25°C for 30 minutes. The pyrite sample was separated from the suspension and dried at room temperature. The captive-bubble method was used for the contact angle measurements [11].

Electrokinetic Measurements Electrophoretic mobilities of the bacterium, obtained from three different culture media, were determined using the Lazer Zee Meter. The pH of the suspensions was measured using Fisher 925 pH meter in conjunction with Fisher glass electrode. The bacterial suspensions were prepared in an electrolyte solution (100 ml of 5 x 10 - 4 M NaC1), and the pH was adjusted using NaOH and HC1. The samples were further conditioned for 15 minutes and the zeta potential values were measured. The reported values are the average of 10 readings.

Adhesion of T. ferrooxidans on Pyrite Surface Adsorption measurements of T. ferrooxidans on pyrite surface were carried out in a 100 ml glass stoppered flask. 100 mg Pyrite sample (74 x 53 #m) was washed with sulfuric acid ( p H = 1.0) for 10 minutes and mixed with 50 ml of T. ferrooxidans (108 cells/ml at pH=2.0). The auspension was stirred for a period of 30 minutes to allow for the adsorption to reach an equilibrium value. The stirring was stopped and the suspension was allowed to settle. The supernatant was withdrawn to determine the equilibrium concentration of the bacterium. The amount of T. ferrooxidans adsorbed on the pyrite surface was determined by the difference in concentration before and after the addition of pyrite. The adsorption density was calculated using the following equation: I" ---- (C 1 - C 2 ) V / W

(1)

where Y is the adsorption density expressed by the number of T. ferrooxidans cells adsorbed per unit weight (g) of pyrite, C 1 and C 2 refer to the bacterial concentration (cells/ml) before and after adsorption, V is the volume of the T. ferrooxidans solution (ml), W is the weight (g) of the pyrite sample used in the experiment. AC Impedance Spectroscopy The passivation or the reactivity of the pyrite mineral was examined using a technique known as AC (Alternate Current) Impedance Spectroscopy. Impedance spectroscopy is a relatively new method of characterizing the electrical properties of materials and their interfaces with electronically conducting electrodes. It uses very small excitation amplitudes which stimulate only minimal perturbation of the test system, thus reducing the errors caused by the measurement technique itself. Further, because the method does not involve a potential scan, measurements can be made in low conductivity solution. In DC (Direct Current) measurements, a special case of AC where the frequency is 0 Hz, the resistance R is defined by Ohm's Law: E = IR

(2)

while in AC, where the frequency is non-zero, the analogous equation is: E = I Z

(3)

where Z is defined as 'impedance', the AC equivalent of resistance. In general, the impedance consists

M. M i s r a et al.

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of a real component Z' and an imaginary component jZ' such that Z = Z' + j Z "

(4)

where 'j' is a constant. In impedance spectroscopy one of the methods to display the Z values makes use of Nyquist plot. In this method the imaginary component of impedance (Z") versus the real component of impedance (Z') is plotted for each excitation frequency as shown in Figure la (data for the equivalent circuit is shown in Figure lb). Impedance measurements were carried out in a frequency response analyzer, which is interfaced with a microcomputer for the data logging and analysis. Schlumberger 1260/Gain Phase Analyzer was used in combination with a Schlumberger 1286 electrochemical interface. The applied frequency range was from 1 mHz to 1 MHz, and the applied potential was 10 mv.

E

cO

U U C c~ U

E E

800 Real Impedance Z ' ( o h m s )

C

I "VV',R1

1t2

Fig. l(a) Nyquist plot and corresponding equivalent circuit (b)

Flotation Experiments The mineral sample was ground to 150 x 53/~m in a ceramic ball mill and dry sieved. Recovery of pyrite was accomplished by a combination of biotreatment and collector addition prior to the flotation. Purified

Effect of growth mediumon pyrite flotation

161

sodium isopropylxanthate was used as a collector molecule and argon gas with a flow rate 104 ml./min. was used in all flotation tests. In order to determine the optimum conditions for the recovery, the operating variables such as pre-washing time of the mineral with dilute sulfuric acid, time and pH of bioconditioning with T. ferrooxidans, collector (xanthate) concentration, flotation time and pH were investigated. The preliminary results have lead to the establishment of optimum conditions which are as follows: (i) prewashing time 10 min., (ii) biotreatment with T. ferrooxidans for a period of 30 min., (iii) xanthate addition (1.6 xl0 - 4 M ) at pH 9 and a conditioning time of 5 min., and (iv) flotation time 90 seconds. A Hallimond tube was used in all flotation tests. The schematic of the flotation test is shown in Figure 2. It must be emphasized that the bioconditioning was carried out at a lower pH 1.8 and collector addition and flotation was performed at higher pH 9.0. The reason being that the bacterial adhesion and its biogeochemical reactions are optimal at lower pH values, whereas xanthate as a collector is effective only at higher pH values. Further, xanthate is believed to decompose in the solutions of low pH values. Flotation experiments were conducted using pyrite and the bacteria harvested from three different culture media.

Mineral

~

pH Conditioning (pH 1.8, Time =10 min.)

Filtration Filtrate

~

Conditioning with T.f. (30 min.)

Filtration

~.j~Conditioning with xanthate ~ ( 5 min. at pH 9.0)

Filtrate

Flotation (90 see.) Tailings (Non floatable pyrite)

Concentrate (Floatable pyrite)

Fig.2 Hallimond tube flotation flowsheet

RESULTS AND DISCUSSION Contact Angles Contact angle of water bubble placed on a polished pyrite surface was measured as a function of T.

ferrooxidans concentration, and the results are plotted in Figure 3. The contact angle of pyrite without T. ferrooxidans was noted to be 49 °. With the adsorption of T. ferrooxidans (at a concentration of 1.9 x 108 cells/ml), the contact angle of pyrite was decreased to 230 indicating the decreased hydrophobicity of mineral. As can be seen, T. ferrooxidans cultured in PDM can make pyrite surface significantly more hydrophilic than that grown in SM.

M. Misra et al.

162 50

45

- e - Pyrite(SM) Pyrite(PDM)

40

35 ~

30

o 25 A v

20

15 0

2

4

6

8

10

12

14

16

18

20

22

T. ferrooxidans Concntration (XIO7 cells/ml) Fig.3 Contact angle as a function of T. ferrooxidans concentration Zeta Potentials

Figure 4 shows the zeta potential of T. ferroxidans cultured in different media as a function of pH. It is evident that the zeta potential values of T. ferroxidans derived from different culture media are different. A greater variation among the bacteria was noted in the PZC values. T. ferroxidans grown in SM, PDM, and PAM were observed to have PZC values at p H = 2 . 0 , p H = 3 . 2 , and p H = 3 . 7 respectively. Similar increases in the PZC values for the bacteria grown on several mineral surfaces have been reported [12]. 10 2 ~-3"L~ 4

5

6

,

7

8

9

10

,

,

,

pH ,-, - I 0 E "d

~

-20

-3o

-40 .It

PM

0

~""-

-50

Fig.4 Zeta potential of T. ferrooxidans clutured in different media as a function of pH

Effect of growth medium on pyrite flotation

163

Adhesion of T. ferroxidans on Pyrite Surface Adhesion of bacteria, grown from three different culture media, was determined on the pyrite surface at pH 2.0. The particle size of the pyrite sample used in these experiments was in the range 75 x 53 #m. The results obtained in this study are shown in Figure 5. It is evident that the growth medium influences the surface properties of bacteria. It is well known that the nutritional composition of the culture medium greatly influences the growth phase [13] and the hydrophobicity of the bacterium [14]. The adsorption of the bacteria cultured in the standard 9K medium was found to be the lowest. In comparison to the standard 9K medium, the adsorption on the other two media viz., PDM and PAM was found to be higher by 80% and 140% respectively. In general, microorganisms are sensitive to the nutritional requirements in their growth phase and starvation brings about changes in the nature of bacteria cell surface and their hydrophobicity [15-17]. In particular, Jerez et al., [18] have reported that T. ferrooxidans undergoes several changes upon phosphate starvation. Amaro et al., [19] reported that phosphate starvation increased the capacity of T. ferrooxidans to attach itself to the sulfide minerals and elemental sulfur. Such an increase was attributed to the changes in the outer membrane proteins and a 25 % increase in the lipopolysaccharides in comparison to the bacteria grown in the standard 9K medium. Phosphate starvation was shown to increase the proportion of high molecular weight proteins with a seemingly corresponding decrease in the low molecular weight fraction [18].

5

-~ 4

53

~9

~2 t9 t~

< °~

0

m

SM

PDM

PAM

Fig.5 Attachment of T. ferrooxidans to pyrite

AC Impedance Spectroscopy of the Sulfur Coated Pyrite In the acidic pH range, the pyrite (FeS2) undergoes an oxidation reaction resulting in the formation of elemental sulfur. The product sulfur, which is coated on the pyrite surface, passivates the mineral and considerably reduces further oxidation. Also a coating of elemental sulfur makes the mineral hydrophobic and enables it to become highly floatable. Figures 6a-b show the results of impedance measurements, as represented by the Nyquist plot, for the mineral pyrite at pH 1.8 in the absence and in the presence of T. ferrooxidans (grown in SM). Figure 6a shows a non-linear increase in the impedance value of pyrite with time indicating that the pyrite is progressively coated with non-conducting sulfur leading to the passivation of mineral. Sulfur deposition is attributed to the superficial oxidation of pyrite mineral and has been well

M. Misra et al.

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documented [20-22]. However, in the presence of T. ferrooxidans the increase in impedance of pyrite was relatively very small (Figure 5b) and can be inferred that the deposited sulfur is further oxidized by the T. ferrooxidans to sulfate thereby making the surface more conducting and less passive. As there is a small net increase in the impedance value of pyrite in the presence of T. ferrooxidans during the 10 day period, it may be concluded that the formation of sulfur is faster than the oxidation of sulfur to sulfate. Sulfur coating of the pyrite makes the mineral more hydrophobic and enhances its floatability. However, on removal or oxidation of coated sulfur, in addition to the attachment of T. ferrooxidans, the pyrite becomes hydrophilic and is depressed in the flotation cell.

-1 700

Without

T.

Ferrooxidans

10

d

I

I

5d

0

-1700 u

With

T.

Ferrooxidans

m

o

I 0

I

I

W

I

I

1

I

1700

Fig.6 Impedance measurement of pyrite (1-10 days), (a) Nyquist plot in the absence of T. ferrooxidans, and (b) Nyquist plot in the presence of T. ferrooxidans

Flotation

Recovery

of Pyrite

The pyrite flotation results obtained with T. ferrooxidans derived from three different culture media are plotted as a function of the bacteria concentration in Figure 7. The conditions of the experiment are described in Figure 2. In each case it was observed that the amount of pyrite reported to the froth decreased with an increase in the bacterium concentration. T. ferrooxidans harvested from different growth media showed large variations in the depression of pyrite and that cultured in the PAM depressed pyrite to the greatest extent in the bacterium concentration range studied.

Effect o f g r o w t h m e d i u m on pyrite flotation

165

100 90

•

SM

O

PDM

•

PAM

80 70 Xanthate: 1.6X10 "4 M 60 50 O

40 30

gt,

20 10 0

I

I

0

2

~

I

I

4

I

I

6

8

~

I

I

I

I

I

I

I

I

10 12 14 16 18 T. ferrooxidans concentration (X107 cells/ml)

20

22

Fig.7 Pyrite flotation recovery as a function of T. ferrooxidans concentration Figure 8 shows the results of pyrite flotation, using T. ferrooxidans grown in PAM and xanthate collector molecule, as a function of bacterial conditioning pH. In all cases the flotation experiments were conducted at pH 9.0. Also included in Figure 8 are the results of pyrite natural flotation unassisted by the biotreatment and collector addition, and the results of pyrite flotation with xanthate as a collector molecule 100

[~-(Z)--------~

~-L

q~

90 Natural flotability

80

X a n t h a t e ( l 0"4M)

70

T f. g r o w n in p y r i t e ( 2 X l 0 s) cells/ml

ID

+ X a n t h a t e ( 1 . 6 X 10"4M)

8 6o = 50 .2 40

•~ 30 20 10 0

J

0

I

1

r

P

2

I

f

I

3

f

4

I

I

5

J

I

6

I

7

pH Fig.8 Flotation of Pyrite as a function of bioconditioning pH--Flotation conducted at constant pH of 9

M. Misra et al.

166

without prior biotreatment with the bacteria. Although flotation in the last two cases was also performed at pH 9.0, the suspensions containing the pyrite were treated in the same manner, that is conditioning at the lower pH values for the same duration of time, as proceeded in the case of pyrite with a combined biotreatment and collector addition. This was done to subject the pyrite surface to the same pH changes and conditioning so that if any surface reactions should occur at low pH values comparisons can still be made for the cases presented in the Figure 8. It can be seen that in the case of pyrite natural flotation most of the pyrite was depressed and only 32 % reported to the froth, which gradually decreased to < 10 % as the conditioning pH was increased. On the other hand, in the case of xanthate alone as a collector more than 93 % pyrite reported to the froth and very little change occured as the pH was varied from 1.8 to 6.5. Finally, in the case of combined bio- and collector treatment of pyrite the recovery was observed to be 43 % at pH 1.2 and decreased slowly till the pH reached 4.0 and decreased further steeply thereafter to 10%. Thus a large decrease in the pyrite recovery, in comparison to the case of xanthate alone treatment, may indicate that the pyrite surface was largely covered by the bacteria and access to the xanthate molecule is prevented.

Mechanisms of Sulfide Mineral Flotation and Depression Mechanism of the xanthate assisted sulfide minerals flotation is well known. It involves the adsorption of xanthate, with a subsequent formation of dixanthogen on the surface of the mineral which increases the hydrophobicity and reports to the froth phase in the flotation cell. However with a prior biotreatment, the depression of pyrite in the flotation cell is believed to be the result of two possible, viz., direct and indirect mechanisms. In the direct mechanism, adhesion of T. ferrooxidans to pyrite surface renders the mineral hydrophilic which causes the depression [23,24]. The adsorption (or adhesion) of bacteria onto the mineral and its pH dependency can be adequately explained with the help of DLVO theory [25]. The indirect mechanism requires no intimate contact or adhesion of the bacteria on the mineral surface. However, it operates through the chemical action of ferric sulphate, a product produced by the bacterial metabolism occurring on the surface of the mineral. Surface oxidation of the pyrite enables it to be hydrophilic, resulting in depression of the mineral. In a recent study Pesic and Kim [26] have reported that surface oxidation at the pyrite mineral surface, caused by T. ferrooxidans over a period of five days, resulted in the formation of a layer of jarosite. The x-ray diffraction method was used to cofirm the formation of jarosite. The product jarosite is formed in an acid producing reaction as shown below: 3 Fe 3+ + X + + 2 HSO 4 - + 6 H20

,~

XFe3(SO4) 2 (OH) 6 + 8 H +

(5)

and has the general formula XFe3(SO4) 2 (OH) 6, where X represents K +, Na +, NH4 +, or H3 O+. Precipitation of jarosite adversely affects bacterial leaching systems due to the formation of impenetrable barrier. However, in the biotreated flotation of pyrite, if jarosite is formed, it may help depress the mineral.

SUMMARY Bioconditioning of pyrite with Thiobacillusferrooxidans produces a significant surface modification of the mineral surface. The flotability of pyrite is significantly reduced, depending on the bacterium concentration, conditioning pH, and the nutritional composition of the culture medium in which the bacterium is grown. Decrease in the pyrite floatability is attributed to the adhesion of T. ferrooxidans on its surface, hydrophobic sulfur oxidation, and the formation of jarosite on the surface of pyrite. From the results obtained in this study the following conclusions were made:

1. T. ferrooxidans, in conjunction with the xanthate collector, can significantly depress pyrite in flotation. 2. The surface properties of the bacterium can be altered by changing the conditions and nutritional

Effect of growth mediumon pyriteflotation

167

composition of the culture medium. The three different media used for the growth of T. ferrooxidans, viz., 9K standard, phosphate-deficient, and pyrite added media have produced bacteria with notable differences in the surface properties, such as contact angles and zeta potentials. The largest change was observed in the bacteria PZC values. 3. In the low pH region, the pyrite surface undergoes superficial oxidation resulting in the formation of an insular layer of elemental sulfur, which increases the impedance of the mineral and also enhances the hydrophobicity. However, in the presence of T. ferrooxidans the elemental sulfur formed on the pyrite surface is further oxidized to sulfate and alters the hydrophilicity/hydrophobicity balance to help depress the mineral. 4. The mechanism of pyrite depression in the flotation cell involves the adsorption of T. ferrooxidans on the mineral surface making it hydrophilic. The adsorption of T. ferrooxidans on pyrite can be explained by the DLVO theory.

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