Letters in Applied Microbiology 1994,18,209-213

Mineral leaching of non-sulphide nickel ores using heterotrophic micro-organisms

P.G. Tzeferis, S. Agatzini and E.T. Nerantzir National Technical University of Athens (NTUA), Department of Mining-Metallurgical Engineering, Laboratory of Metallurgy, Athens, Greece

GWG/219: accepted 31 October 1993

P.G. TZEFERIS, s. AGATZINI AND E.T. NERANTZIS 1994. Laboratory studies were conducted on microbial leaching of non-sulphide nickel ores not amenable to conventional mineral processing operations. The results showed that extensive low-grade laterite domestic sources are generally amenable to bioleaching when micro-organisms were cultivated in the presence of the ore. Nickel recoveries were as high as 60% using hydroxycarboxylic acid producing strains of Aspergillus and Penicillium codes A3, P2. Cobalt recovery achieved was around 50%. Losses of soluble nickel in the fungal biomass were found to be 3.5-10.8%. Chemical analysis of the leach liquors showed the presence of significant amounts of citric, oxalic and other organic acids, indicating that leaching may be ascribed to the production of these metabolic products of fungal activity.

INTRODUCTION

Nickel is of vital importance in many industrial processes. Nickel ore deposits are found either as sulphides or as later- ites. The dominating importance of lateritic ores for the future supply of nickel becomes obvious when one con- siders that about 80% of the presently known nickel reserves are associated with lateritic types of ore (Blanco and Holliday 1981 ; Simons 1988). As sulphide reserves are exhausted, there will clearly be a need for technological breakthroughs in extraction of nickel from laterites.

Nickel from laterites is produced both pyrometallurgi- ally and hydrometallurgically. Most of current industrial practice, however, is either energy-intensive or presents severe engineering problems. A microbial based process could possibly provide an economical, more competitive and environmentally safer alternative to the existing pro- cesses.

A number of workers have demonstrated the mineral leaching ability of Aspergillus, Penicillium and other aci- dophilic fungi, which are known as efficient carboxylic acid producers. The results reported so far have been concerned with the dissolution of aluminium from bauxides (Groudev and Gencev 1978) and aluminium bearing rocks (Groudev and Groudeva 1986), magnesium from manganese ores (Mercz and Madwick 1982), potassium from laucite (Rossi 1978), titanium from granitic rocks (Silverman and Munoz

Correspondence t o : Dr Peter Tztfmis, 45 Gythiou str., 185 44 Piraeus, Greece.

1970, 1971), copper and zinc from various oxidized ores (Dave et al. 1981) and nickel from laterites not amenable to upgrading by means of the usual conventional operations (Bosecker 1985).

In the present study, a research programme was under- taken to identify micro-organisms, media and biological mechanisms which could extract, efficiently and eco- nomically, nickel and cobalt from extensive Greek low- grade laterite deposits. A series of chemical leaching experiments had been previously conducted to evaluate whether organic acids produced in the metabolism of car- bohydrate media by fermentation pathways were capable of solubilizing nickel from laterites (Tzeferis 1991).

The work was done a t the Laboratory of Metallurgy of the National Technical University of Athens (NTUA) in cooperation with the Centre for Biotechnology and the Mineral Resources Engineering Department of the Imperial College, London.

MATERIALS AND METHODS

Ore samples

Lateritic nickel ore samples were obtained from deposit ‘Litharakia’ (Euboea island, Greece) and ‘Kastoria’ (Kastoria, Northern Greece). Investigation by X-ray dif- fraction and electron microprobe analysis revealed that the

210 P.G. TZEFERIS ET A L .

'Litharakia' sample was the 'typical low grade' limonitic lat- erite with 0.73% Ni mostly in chlorite form while the 'Kas- toria' sample was of the garnieritic type with relatively high nickel and magnesium contents, 4.23% and 10.17% respec- tively. All ore samples were crushed and ground to produce particle sizes of less than 150 mesh (106 pm).

Micro-organisms

One Penicillium strain (code F1) was isolated from an actively laterizing Greek ore. Various strains of Aspergillus (codes Al, A2, A3) and Penicillium sp. (codes P2, P14, P24, P6) were also provided by Greek, English and German col- lections, with characteristics likely to be beneficial to the leaching process. All strains were tested for their ability to: (i) produce organic chelating acids; and (ii) tolerate high nickel and hydrogen ion concentrations as in an actual leaching process.

The following substrates were used for growth and bio- leaching experiments :

A Glucose medium was used for both growth and screening leaching experiments. A Sucrose medium was chosen from literature (Shu and Johnson 1947, 1948; King 1985) and optimized by means of a factorial analysis to maximize the yield of citric acid (Tzeferis 1991). Industrial grade Greek beet molasses (sucrose 47%) were also used in an effort to reduce the overall cost of bioleaching process. _ _

All feed reagents were laboratory Analar grade and after preparation were sterilized at 121°C and 15 psi for 20 min. The formulae for media used in bioleaching experiments are given in Table 1.

Bioieaching experiments

The leaching tests were carried out in conical flasks con- taining known weights of laterite ore and volumes of culture medium. The flasks containing the nutrient-mineral systems were sterilized by autoclaving to prevent inter- ference by indigenous organisms. The organisms, pre- viously acclimatized to grow in the presence of the ore and dissolved nickel ion, were cultivated in a mixture of the nutrient and finely ground ore. For inoculation, a 10% v/v of fungal spore suspension was added. Inoculated flasks were incubated at 30"C, 400 rev min-', in a magnetically stirred thermostatic bath IKA-RTM 9. Samples were removed at regular intervals and metal concentrations in the supernatant were measured by atomic absorption spec- trophotometry (AAS).

At the end of the experiments the solid leach residues, having been separated from the liquid, consisted of laterite and fungal biomass since it was practically impossible to

Table 1 Composition of microbiological media used

Glucose Sucrose Molasses Constituents medium (g 1-')

Glucose 150.0 Sucrose 150.0 Molasses 150.0 NaNO, 3.0 (NH,),CO, 1.8 (NH,),HPO* 1 *6 K2Hm4 1 *o 0.25 MgSO, .7H,O 0.5 1.2 KCI 0.5 FeSO, . 7H,O 0.01 H + (PH) 4.3 5.0 Fez+ 0.01 mg I - ' Zn2+ 0.01 mg I - ' cu2+ 0.01 mg I - ' 1.3% Water 1.0 I 1.0 1 1.0 1

separate the biomass from the residual mineral powder. The solids were leached with hot water in successive stages, until no nickel was obtained in solution. It was then assumed that the remaining nickel was that taken up by fungal biomass. The solid residues were heated to 700°C in a platina crucible to decompose the biomass and liberate its metal content as a residual ash. This operation was fol- lowed by leaching with slightly acid water at 90°C and the filtrate was analysed for nickel content by AAS. Nickel in residues could not be detected by XRD methods as its content was too low. Citric and oxalic acids produced by fungal metabolism were determined using biochemical ana- lytical methods (Boehringer Manual 1989).

RESULTS AND DISCUSSION

All strains tested, when acclimatized, exhibited a much improved tolerance, especially the Penicillium sp. which were less effected by nickel than the Aspergillus sp. However, most of the fungal strains could hardly grow in a laterite pulp of 10% density, even after their adaptation to nickel solutions. A procedure of progressive acclimatization was thus undertaken to adapt the strains to laterite ore by increasing its content in the culture medium. Table 2 shows the growth rates of fungal mycelium when grown on agar plates with 10% lateritic ore incorporated in sucrose media.

Figure 1 shows the nickel recoveries achieved during bioleaching of the low grade ore in the presence of various organisms in sucrose medium for over 48 d. The most effective strains, P2 and A3, could extract nickel up to 60% from the ore. Losses of dissolved nickel in the fungal biomass were found to be 3.5% for strain P2 and 10.8% for

LATER I TE H ETE R OTR 0 P H I C LEACH I N 0 211

Table 2 Adaptation in laterite environment

Organism Al A2 A3 P2 P6 P14 P24 F1

Growth rate before serial 0.17 0.29 0.18 0.24 0.27 0.28 0.15 0.14

Growth rate after serial 0-6 1 0-46 0.82 1.02 0.78 0.63 0.95 0.70 acclimatization process (cm d- ')

acclimatization process (cm d - ')

Growth medium: 15% sucrose plus 10% laterite ore.

strain A3. Cobalt extraction was about 50%. Maximum dissolution of iron was around 25% and coincided with an increased production of oxalic acid.

The mechanism of fungal leaching appeared to be one in which the successful micro-organisms produced adequate amounts of citric acid and other organic metabolites, which then attacked the laterite ore. As shown in Table 3, high levels of nickel extraction corresponded to high citric acid

Leaching temperature: 30°C Agitotion speed: 400 rew min-' Pulp density: 10% Initial pH: 4.3 - ae 50 'I

0 10 20 30 40 L

50 I

Time ( d )

Fig. 1 Recovery of nickel as a function of time during leaching of low grade ore by various fungi in sucrose medium. 0, P2; 0, A3; 0, F1; 0, Al; A, A2; A, P6; 0, P24; V, P14; x , control

Table 3 Citric acid production us nickel extraction

Citric acid concentration (g 1 - ')

In In fermented Nickel bioleaching sucrose recovery

Micro-organism solution media (%I P2 57.2 21.0 61.5 f 5.35 A3 61.5 42.0 52.8 f 4.74 P6 20.4 3.5 24.6 F1 0.0 0.0 5.75 A2 8.4 3.5 12.8

production determined in the leach solution. The organisms, as shown in the same table, produced much more citric acid in the presence of the ore than in the ore- free fermentation medium. This behaviour was attributed to an induction mechanism caused at the vicinity of the hyphae with the laterite particles. It seems that the compli- cated and possibly toxic system 'micro-organism-nutrient substrateore' could be positive for nickel leaching. It is, however, possible that other than citric acid metabolites (oxalic and other minor components) might have played a role as chemical leaching of the ore by synthetic solutions of the same citric acid concentrations as the above gave lower nickel recoveries than those obtained during bio- leaching (Tzeferis 1991).

When sucrose was replaced as the carbon source by glucose, the results of microbial leaching tests with all strains were in the range 5 2 0 % (Figs 2 and 3). Both Aspergillus and Penicillium sp. grew well in this medium together with nickel and laterite but did not produce sig- nificant quantities of citric acid, despite the rapid reduction of pH. This low selectivity of glucose media used for citric

Leaching temperature: 30°C Pulp density: 10% Agitation speed: 400 rev min-'

I 0- I0 20 30 40

Time ( d )

FIg. 2 Recovery of nickel as a function of time during leaching of high grade ore by various fungi in glucose medium. For definition of symbols, see Fig. 1 legend

212 P . G . TZEFERIS ET A L

trace metals and reduce the ash content, thereby resulting in improved nickel yields during leaching. Leaching temperature: 3OoC

Pulp density: 10% Agitation speed: 400 rev min-'

CONCLUSION

Greek nickeliferous laterites, although mineralogically vari-

Time ( d )

Flg. 3 Recovery of nickel as a function of time during leaching of low grade ore by various fungi in glucose medium. For definition of symbols, see Fig. 1 legend

production could be an explanation for the low nickel extraction obtained in this medium.

Unrefined Greek beet molasses, kindly provided by the Hellenic Sugar Industry SA, were also used in bioleaching experiments as an alternative and more economical source of carbon and energy. The nickel recoveries, shown in Fig. 4, were generally lower than in the sucrose medium, pos- sibly due to high ash (11.4%) and metals content (K: 3*5%, Na: 1*7%, Fe: 1.3%) in this medium. Research in progress, including treatment of molasses with potassium ferrocyanide, will possibly enable the removal of deleterious

40 Leaching temperature: 3OoC

Agitation speed: 400 rev min-'

z. 25 - a-"

ii

-

0- 10 20 30 40 50

Time ( d )

Fig. 4 Recovery of nickel as a function of time during leaching of low grade ore by various fungi in molasses medium. 0, P2, 0, A3; 0, A2; 0, F1; A, A l ; A, P6; V, P24; x , control

able and complex, were generally amenable to laboratory bioleaching techniques, in which the medium and inocolum were added to the ore in flasks. Maximum yields obtained were up to 60% of nickel, 50-55% of cobalt and 20-25% of iron codissolution. It is believed that nickel and cobalt were extracted mainly by citric acid and other substances produced by fungal metabolism, whereas iron was dissolved by oxalic acid, also determined in the leach liquor.

ACKNOWLEDGEMENTS

Financial support from the Commission for European Communities, DGXII, the Greek General Secretariat of Research and Technology of Greece and the General Mining and Metallurgical SA LARCO is gratefully acknowledged. The authors would like to thank Dr D. Leak, Dr K. Alibhai and Dr A. Dudeney, Imperial College of London, for their helpful collaboration in this research project.

REFERENCES

Blanco, J.L. and Holliday, B.M. (1981) Energy swings at nickel. Journal of Metals April, 50-51.

Boehringer Mannheim GmbH User's Manual (1989) Methods of Biochemical Analysis and Food Analysis using Test-Combinations. Revised edition, pp. 142.

Bosecker, K. (1985) Leaching of lateritic nickel ores with heter+ trophic microorganisms. In Proceedings of the 6th International Symposium on Biohydrometallurgy, Vancouver, Canada, August

Dave, S.R., Natarajan, K.R. and Bhat, J.V. (1981) Leaching of copper and zinc from oxidized ores by fungi. Hydrometallurgy 7,235-242.

Groudev, S.N. and Gencev, F.N. (1978) Bioleaching of bauxites by wild and laboratory bred microbial strains, 4th Congress I.C.S.O. B.A, Athens, 1, pp. 271-278.

Groudev, S.N. and Groudeva, V.I. (1986) Biological leaching of aluminium from clays. In Workshop on Biotechnology for the Mining, Metal-rejning and Fossil Fuel Processing Industries ed. Ehrlich, H.L. and Holmes, D.S. Biotechnology and Bioengi- neering, Symposium No. 16, pp. 91-96.

King, A.B. (1985) The biodegradation of nepheline by Aspergillus niger. PhD Thesis, University of London, 328 pp.

Mercz, T.I. and Madwick, J.C. (1982) Enhancement of bacterial manganese leaching by microalgal growth products. Proceedings of the Australasian Institute of Mining and Metallurgy, No. 283, September, pp. 43-46.

21-24, pp. 367-382.

LATERITE HETEROTROPHIC LEACHING 213

Rossi, G. (1978) Potassium recovery through leucite bioleaching : possibilities and limitations. In Metallurgical Applications of Bacterial Leaching and Related Microbiological Phenomena ed. Mum et al. pp. 297-319. NY, San Francisco, London: Aca- demic Press.

Shu, P. and Johnson, M.J. (1947) Effect of the composition of the sporulation medium on citric acid production by A. niger in submerged culture. Journal of Bacteriology 54, 161-167.

Shu, P. and Johnson, M.J. (1948) The interdependence of medium constituents in citric acid production by submerged fermentation. Journal of Bacteriology 56, 577-585.

Silverman, M. and Munoz, E. (1970) Fungal attack on rock: solu- bilization and altered infrared spectra. Science 169,985-987.

Silverman, M. and Munoz, E. (1971) Fungal leaching of titanium from rock. Applied Microbiology 22, 923-924.

Simons, C.S. (1988) The production of nickel: extractive metallurgy-past, present and future, In Metallurgy of Nickel and Cobalt, Proceedings of a Symposium-1 17th TMS Annual Meeting, Phoenix, Arizona, January 25-28, pp. 91-134.

Tzeferis, P.G. (1991) Bioleaching of nonaulphide nickeliferous ores using heterotrophic microorganisms. PhD Thesis, National Technical University of Athens, Greece, pp. 376.