Study on reaction mechanism of bioleaching of nickel and cobalt from lateritic chromite overburdens

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International Biodeterioration & Biodegradation 65 (2011) 1035e1042

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International Biodeterioration & Biodegradation

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Study on reaction mechanism of bioleaching of nickel and cobalt from lateriticchromite overburdens

S.K. Behera, P.P. Panda, S. Singh, N. Pradhan, L.B. Sukla*, B.K. MishraInstitute of Minerals & Materials Technology (CSIR), Bhubaneswar 751 013, India

a r t i c l e i n f o

Article history:Received 25 February 2011Received in revised form21 July 2011Accepted 5 August 2011Available online 15 September 2011

Keywords:Chromite overburdensAspergillus nigerBioleachingNickelTEMHPLCOxalic acidReaction kinetics

* Corresponding author. Tel.: þ91 674 2581635; faxE-mail address: [email protected] (L.B. Sukla).

0964-8305/$ e see front matter � 2011 Elsevier Ltd.doi:10.1016/j.ibiod.2011.08.004

a b s t r a c t

Depletion of high-grade ores and presence of significant quantities of metals in low-grade oxide ores hasenforced to utilize the overburdens (COB) and wastes (low-grade ores) generated during mining oper-ations. The impact of ore mineralogy and mineralemicrobe interaction during bioleaching could not beignored. Seeking to the need, a systematic study was performed to establish the reaction mechanisminvolved for recovery of nickel and cobalt from chromite overburden (COB), Sukinda, Orissa using pureculture of Aspergillus niger. Mineralogical analysis reveals a complete conversion of goethite intohematite phase leading to exposure of nickel particles into the micro-pores and cracks developed in thematrix which was initially found to be intertwined in the goethite lattice. As a result, it became moresusceptible to attack by the fungal bio acids which in turn accelerate the dissolution rate. Organic acidslike oxalic and citric acids were detected in the culture filtrate using HPLC. TEM analysis of the leachedsamples shows that nickel dissolute into the solution leaving a porous space in the matrix of the hematiteby forming nickel oxalate or nickel citrate. Kinetics of the nickel bioleaching was studied to support themechanism of the reaction. It was observed that the initial rate of reaction follows the chemical controldissolution reaction where as the later part fits to shrinking core model. 18% of nickel and 37.8% of cobaltwas recovered from pre-treated COB at 2.5% pulp-density with 10% (v/v) fungal inoculum at 30 �C within25 days in shake flask while 32.5% of nickel and 86% of cobalt was recovered in bioreactor.

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1. Introduction

Laterites are oxidic ores widely distributed in the tropicalregions, formed due to laterization, a weathering process of ultra-mafic rocks favoured by warm climate and abundant rainfall.Sulphides and laterites are two forms of nickel ore occur in nature, ofwhich the sulphides are the major source for nickel extraction. InIndia, lateritic chromite overburdens are generated at Sukindamines near Jajpur, Orissa located at 20�580000N 85�550000E occupyingan important geographical position in mineralogical map of India.However, it has been estimated that the lateritic ores constitutearound 85% of known nickel reserves, a significant future source ofnickel (Mohapatra et al., 2007). Due to stringent environmental lawsand high demand has forced the use of low-grade oxidic ores as analternative for nickel extraction. It has been estimated that Sukindachromite overburden contains 0.8e1.0% of nickel and 0.03e0.04% ofcobalt (Sukla and Das, 1987) which needs to be exploited.

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The existing hydrometallurgical processes for leaching of nickellaterite such as high pressure acid leaching, Caron process, ferro-nickel and nickel matte smelting techniques are energy intensiveand the operational costs are high (Tang and Valix, 2006). This hasnecessitated the development of feasible processes that couldaddress the economic and environmental limitations in the afore-said techniques. In this regard, microbial mineral processing routescould potentially meet the challenges in processing of nickellaterites. Secondly, it could provide an alternative technology forprocessing abundant reserves of low-grade laterite nickel ore.

Literature survey shows extraction of nickel from lateritic oreusing microbial processes (Bosecker, 1985; Alibhai et al., 1992;Sukla et al., 1993; Tzeferis, 1994; Valix et al., 2001). Unlike othermicro organisms, fungus solubilizes metals from lateritic ore by theformation of organic acids viz. citric acid, oxalic acid, gluconic acidetc. via acidolysis, complexation and chelate formation (Burgstallerand Schinner, 1993a; Bosecker, 1997; Brandl, 2001; Tsekova et al.,2010). Detail mechanism of heterotrophic bioleaching is stillunder debate. Therefore, proper investigation is required toestablish the reaction mechanism by identifying and quantifyingdifferent types of organic acids produced during the process.

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Impact of mineralogy on bioleaching cannot be ignored. In thelateritic ore, nickel occurs in an absorbed state within the goethitematrix along with other mineral phases. Due to low solubility andcomplex structure extraction of nickel from the goethite matrix isvery difficult. To overcome from this problem the COB wassubjected to pre-treatment process at 600 �C under normal atmo-spheric conditions. As a result of thermal activation, phase trans-formation occurs by dehydroxylation of goethite matrix in the rawCOB (Landers and Gilkes, 2007; Jinhui et al., 2009). It has beenreported that due to thermal pre-treatment process nickel & cobaltrecovery was enhanced through bioleaching (Ruan et al., 2002;Valix and Cheung, 2002).

Addressing the need, the present study deals with enhancementand establishment of reaction mechanism for nickel and cobaltrecovery through pre-treatment of COB using pure strain ofAspergillus niger in shake flask and 40 L stainless steel bioreactor.

2. Materials and methods

2.1. Chemical and mineralogical analysis of COB

Samples were collected from Sukinda Mines, Orissa, India. Thesamples were crushed and sieved to obtain a particle sizeof �75 mm. The samples were air dried and the metal concentra-tions were determined in atomic absorption spectrophotometer(AAS) after digesting in concentrated HCl. Mineralogical analysis ofthe original and leached samples were carried out by means ofPhillips Diffractometer (PW3710) with a radiation operating at40 kV and 30 mA to identify major and minor minerals.

Experiments were performed using two different forms of theCOB, designated as raw and pre-treated COB. The pre-treatment ofCOB was done in a muffle furnace operated at 600 �C for 5 h undernormal atmospheric conditions.

2.2. SEM and TEM analysis

Scanning electron microscopy (SEM) was carried out on raw andpre-treated COB samples with a size range of 5� 5� 5mmwith ultrathin film of gold coating using an ion sputter JFC- 1100 and studiedunder a Japanese make electron microscope (JEOL-JXA-8100). Theworking heightwasmaintained at 15mmwith a voltage ranging from10kVto25kV.Polishedsamplesobtainedpriorandaftercompletionofthe experiments, were vacuum dried using a vacuum coater. Thesamples for the transmission electron microscope (TEM) investiga-tionswere imaged in (FEI, TECNAI G2) 20 (Netherland), equippedwithenergy dispersive X-ray spectroscopy (EDX) operating at 100 kV.Samplesweredispersed inethanol andsubjected toultrasonication for5min30 ml of the sampleswere coated to the carbonecoated gold gridand dried before being inserted into the column of the TEM chamber.

2.3. Micro organism and growth conditions

Pure culture of A. niger (MTCC No:- 6997) was used for the study.The fungal strain was isolated from COB of Sukinda mines andgrown in Bromofield medium containing (g/L): MgSO4 7H2O - 0.75,KH2PO4 - 0.25, (NH4)2SO4 - 0.25, yeast extract- 1 and sucrose-20, pH6.8 at 30 �C. Isolates were identified by their colony characteristics,spores morphology and microscopic observations. Spore suspen-sion (106spores/ml) of 5e7 days old culture was used as inoculumfor further experimentation.

Culturefiltrateswere collected before, after andduring bioleachingprocess andwere analyzed by HPLC to quantify the amount of organicacidsproduced. Foranalysis,ModelAgilent-1100,Agilent technologies,Waldbronn Analytical Division, Germany) containing Zorbax EclipseXDB-C18 column (150 � 4.6 mm, i.d) was used. Two mobile phases

(A&B)were involved in theHPLC analysis. Themobile phase usedwasmethanol: water (30:70 v/v) with a flow rate of 1.2 ml/min withaquaternarypump(ModelNo-G1311A)operatedat60 �Cwith thermostart (Model No -G1316A). Solvent A was methanol (HPLC grade) andsolvent B was Millipore water containing 0.01 mol/l of KH2PO4 solu-tion, pH was adjusted to 2.32 using phosphoric acid. HPLC gradestandardsofoxalic acidandcitric acidwereused. 20ml of sampleswereinjected manually in the injection port along with the standard acidsolutions. The organic acid levels were quantified by UV detection at210 nmwith diode array detector (DAD, Model -G 1315A).

2.4. Bioleaching assay

2.4.1. Shake flask experimentsTwo set of leaching experiments were carried out at 2.5% pulp-

density (w/v) using raw and pre-treated COB in 250 ml Erlenmeyerflasks containing 100 ml of Bromofield medium. Initial pH of themediumwas 6.8. Each flask was inoculated with 10 ml (106spores/ml) of spore suspension and incubated for 25 days at 30 �C ona rotary shaker at 150 rpm. Samples were drawn at regular intervalsand were analyzed by AAS to determine the percentage of nickeland cobalt leached in the liquidmedium. Control experiments werealso performed without addition of micro organisms. Asepticconditions were maintained throughout the experiment.

2.4.2. Bioreactor leaching2.4.2.1. Bioreactor description. Experiments were conducted ina 40 L single-stage stainless steel (SS-316) bioreactor equippedwithstirrer and air sparger. The schematic diagram of the bioreactor isshown in (Fig. 1). The bioreactor is designed with a flat top andhemispherical bottom with a maximum solideliquid ratio of 40%.The pulps are stirredwith amechanical stirrer having air dispersingblades rotating at variable speeds. The flow of air is controlled by anairflow meter and the agitation speed by a tachometer. An elec-tronic control panel is attached to the bioreactor for monitoringtemperature, pH, Eh.

The experiment was performed with pre-treated COB at pulp-density of 2.5% (w/v), with an initial pH of 6.8 at 30 �C, agitationspeed-150 rpm, flow rate-2 l/min and inoculum concentration-106

spores/ml for incubation period of 25 days. Slurry samples from thereactor were collected at regular interval for the analysis of nickeland cobalt by AAS.

3. Results and discussion

3.1. Characterization of COB

The detail chemical analysis of COB shows that, the raw COBcontains 0.99% nickel, 0.03% cobalt, 48.88% iron, 2.59% chromiumand 0.21% manganese, whereas after pre-treatment of COB 1.02%nickel, 0.04% cobalt, 50.85% iron, 3.65% chromium and 0.35%manganese were detected. The composition map of COB sampleshows that nickel occurs in an absorbed state within the goethitematrix (Fig. 2a). Pre-treatment of COB results in complete conver-sion of goethite into hematite, subsequently leading to homoge-neous distribution nickel particles throughout the matrix (Fig. 2b).Comparison between chemical analysis of raw and pre-treated COBshowed an increase in total nickel and iron content. The surfacearea of the ore particle increases due to pre-treatment of COB, theinitial surface area was 23.2 m2/g and after pre-treatment thesurface area of COB particle changes to 45.1 m2/g. X-ray diffractionspectrum (Fig. 3) of the pre-treated COB indicates the peaks asso-ciated with goethite which was originally present in the raw orehad almost disappeared and there was also a marked increase inintensity of peak bands of hematite. As a result development of

Fig. 1. Schematic diagram of bioreactor SS- 316.

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micro-pores and cracks in the particles occurs which leads to betterpercolation of lixiviate into the matrix.

3.2. Characterization of the isolates

The isolate was identified as A. niger. Microscopically the colo-nies consist of a compact white basal felt covered by a dense layer ofblack conidial heads. Conidial heads are large in diameter, globose,dark brown, becoming radiate and tending to split into severalloose columns with age. Conidiophores are smooth-walled, hyalineor turning dark towards the vesicle. Conidial heads are biseriate.Conidia vary from globose to subglobose, dark brown to black andare rough-walled.

3.3. TEM analysis

TEM analyses were carried out with the samples collectedbefore and after leaching. It was observed that due to surfacediffusion, appearances of circular crystals with rounded ends occur

containing hematite (Fig. 4a). This rounded ends of the circularcrystals were more pronounced prior to leaching. These crystalsvary from 90 to 150 nm in length and 6e15 nm in diameter. Thesefindings were similar with Schwertmann and Latham (1986) andLanders and Gilkes, (2007). TEM analysis of the leached samplesshows that during progressive leaching, nickel dissolute into thesolution leaving a porous space in the matrix. (Fig. 4b) depicts thepartial release of the nickel from the hematite after leaching andresults in the formation of irregular pores throughout the matrix.Shape of the pores varies from spherical to sub-spherical while insome cases they merged to form larger pores. The size of the poresvaries from 10 to 30 nm. Similar results were also reported byPomies et al. (1999) and Löffler and Mader (2006).

3.4. Quantification of organic acids using HPLC

Organic acids such as oxalic acid, citric acid, gluconic acid etc areexcreted into the medium as metabolic products and subsequentlydissolve heavy metals by forming salts and chelates (Valix et al.,

Fig. 2. (a & b): Composition map showing (a) Nickel, manganese and cobalt occur in absorbed state with Fe in raw COB. (b) Nickel and cobalt is found expelled from goethite matrixafter pre-treated COB.

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2001). During metabolism, fungus converts sucrose or othercarbohydrates into variety of products including organic acidswhich leads to lowering of pH. The accumulation of organic acids bythe microbes results in drop of pH. It was observed that there wasgradual decrease in pH of the leaching system from 6.8 to 2.8 due tothe formation of organic acids. HPLC results showed the presence ofboth oxalic and citric acid. Initially citric acid was detected in theculture filtrate but after the 4th day, presence of oxalic acid wasmore pronounced.

Acid production:

Glucose=Sucrose��!Fungus

Oxalic=Citric=Gluconic acid (1)

3.5. Possible mechanism

As reported, heterotrophic micro organisms are able to mobilizemetals by (i) formation of organic acid (ii) oxidationereductionreactions (iii) extraction by complexity agents (iv) chelate

Fig. 3. X-ray diffraction spectra o

formation. Bioleaching processes are mediated due to the chemicalattack by the excreted organic acids by the micro organisms on tothe ore. Therefore acid production by fungus will be helpful inleaching of metals from ores. The acids usually have dual effect ofincreasing metal dissolution by lowering the pH and increasing theload of soluble metals by complexing/chelating into solubleorgano-metallic complexes (Burgstaller and Schinner, 1993b). Inthis case the organic acids act as chelating agents forming nickel eoxalate and cobalt e oxalate. Oxalic acid contains two carboxylgroups, so the possible complexes of nickel cation with oxalateanion are expressed as:

ðiÞC2H2O4 4 ðC2HO4Þ1�þHþ ðpKa1 ¼ 1:27ÞðC2HO4Þ1�þNi2þ 4 NiðC2HO4Þ2

ðNickel oxalate complexÞ(2)

ðiiÞðC2HO4Þ1� 4 ðC2O4Þ þHþ ðpKa2 ¼ 4:20ÞðC2O4Þ2�þNi2þ 4 NiðC2O4ÞðNickel oxalate complexÞ

(3)

f raw and pre-treated COB.

Fig. 4. (a & b): TEM analysis of pre-treated COB (a) Before leaching (b) After leaching.

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Similarly, citric acid contains three carboxyl groups. When it isfully dissociated, so the possible complexes of nickel cation withcitrate anion are expressed as:

C6H8O7 4 ðC6H5O7Þ3�þ3Hþ ðpKa3 ¼ 6:39Þ2ðC6H5O7Þ3�þ3Ni2þ 4 Ni3ðC6H5O7Þ2

ðNickel citrate complexÞ(4)

Previous literature on the subject suggested that the leachingefficiency of heterotrophic micro organisms depends on the extentof the production of organic metabolites, which are excreted in tothe culture medium and lower the pH of medium. This induces thedissolution of metals into the solution. This fact also hold true withour findings where A. niger with maximum acid and biomassproduction shows maximum solubilization.

3.6. Shake flask study

Fig. 5 shows extraction of nickel and cobalt from pre-treatedCOB as a function of time. It was found out that a maximum 18%of nickel and 37.8% of cobalt was recovered from pre-treated COB at2.5% pulp-density with 10% (v/v) fungal inoculum at 30 �C forduration of 25 days.1.76% of nickel and 5.3% of cobalt was recovered

Fig. 5. Plot showing % of metal extracted versus pH as a function of time from raw andpre-treated COB using Aspergillus niger in shake flask (Conditions: PD:2.5% (w/v),Temp.: 30 �C, Inoculum size: 10%(v/v), Sucrose Conc.: 2%, Duration: 25 days, 150 rpm).

from raw COB under above mention conditions. This was due to thefact that nickel present in the raw COB was tightly entangled in thelattice and therefore not easily dissolved and less susceptible toattack by the micro organisms, hence low recovery was observed inraw COB. While in pre-treated COB higher recovery rate was ach-ieved due to conversion of goethite into hematite as a result ofwhich nickel is exposed to the micro-pores and cracks developed inthe particles at higher temperature.

It was observed that during progressive leaching, pH of themedium decreases from 6.8 to 3.6 (Fig. 5) indicating completeconsumption of sucrose in medium leading to production oforganic acids. Quantification of organic acids was done using HPLC.It was found out that 0.441 mol/l of oxalic acid and 0.112 mol/l ofcitric acid was produced during the leaching period respectively.Apart from organic acid production about 16 gm/100 ml wetbiomass was generated within 25 days. Control set up experimentsshows negligible recovery.

3.7. Bioreactor leaching

Bioreactor leaching consists of three phases such as solid phase,an aqueous phase and a gaseous phasewhich is amixture of oxygenand carbon dioxide. Residence time, flow rate and recirculationtime of the slurry can be adjusted and controlled. Thorough mixingof the three phases is essential for the effective encounter of solidparticles with the micro organisms as well as chemically activemolecules thus helps in the extraction process. Hence, experimentswere conducted at 2.5% (w/v) pulp density with 10% (v/v) fungalinoculum pH- 6.8 at 150 rpm with flow rate of 2 l/min.

Fig. 6 shows plot of nickel and cobalt recovery versus time.A maximum 32.5% of nickel and 86% of cobalt was recovered frompre-treated COB within 25 days of progressive leaching. Higherextraction rate of nickel and cobalt from pre-treated COB was dueto pre-treatment process of COB at 600 �C which results incomplete conversion of goethite into hematite phase. Thisincreased leach-ability may be attributed to the increased porosity,increased surface area and change in reactivity of the COB due toextensive dehydration and dehydroxylation of various hydratedoxideminerals. Fig. 8 shows a gradual decrease in pH from 6.8 to 2.8due to production of organic acids by A. niger. Due to economicalpoint of view low concentration of sucrose was taken for theexperimentation. Initially the sucrose concentration in mediumwas 2% but after 9th day of progressive leaching almost all sucrosewas consumed therefore again 2% of sucrose was added (Fig. 7) tothe medium for continuation of the process. The sucrose in the

Fig. 8. (a & b): Graph showing kinetics of Nickel extraction from pre-treated COB inBioreactor (a) Application of Eq. (2) for initial rate of extraction (b) Application of Eq.(3), for overall rate of extraction.

Fig. 6. Plot showing % of metal extracted versus pH as a function of time from pre-treated COB using Aspergillus niger in bioreactor (Conditions: PD:2.5% (w/v), Temp.:30 �C, Inoculum size: 10%(v/v), Duration: 25 days, Agitation speed: 150 rpm, Aeration:2 l/min).

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media was carbon source which converts to the organic acids(leaching agents) by the fungus during its cellular metabolism.During the study it was observed that the rate of consumption ofthe sucrose in bioreactor wasmore in compare to that in shake flaskscale. The higher rate of the sucrose consumption in bioreactor iscomplimented with the higher rate of growth of the fungus. Thegrowth conditions for the fungus in bioreactor are more supportivethan shake flask might be due to higher surface area, aeration,maintenance of the optimum temperature and agitation of thereaction mixture. Higher growth rate of fungus leads to maximumproduction of organic acid in the leaching system resulting inhigher nickel and cobalt recovery. To fulfil the need sucrose wasadded at regular intervals.

Biomass generated was 22 g/100 ml. As describe earlier inSection 3.3, the amount organic acids produced by A. niger wasestimated by HPLC. Organic acid production starts after a lag phaseand reached to maximum at the onset of stationary phase. Initially0.215 mol/l of citric acid was detected in the leaching system but on9th day, production of oxalic acid was more pronounced than citricacid. Around 1.172 mol/l of oxalic acid was produced within 25thdays. Prolonged leaching leads to decrease in production of organicacids in the leaching system, which might be due to the decreasedavailability of nitrogen source in medium, the age of fungus anddepletion of sugar contents and rise in the pH of the system. Similartype of work has also been reported by Wieczorek and Brauer in

Fig. 7. Plot showing pH versus sucrose concentration as a function of time from pre-treated COB using Aspergillus niger in bioreactor.

1998. Therefore seeking to the need addition of extra sucrose (2%)to the leaching system became necessary to maintain the growth ofthe fungus. A general observation was that only carbon sourcesallow fast growth of A. niger which leads to carbon metabolism.Sucrose concentration is directly proportional to the growth of thefungus and had an adverse effect on the enzyme secretions andorganic acid production. Hossain et al. in 1984 reported that highconcentration of appropriate carbon sources leads to repression ofa-ketoglutarate dehydrogenase. Thus, the effect and need foraddition of carbon source becomes essential in terms of enzymerepression which holds true in our case. Samples were examinedunder the light of phase contrast microscope and were observedthat the size of the mycelia and their shape reduces as the sucroselevel goes down. Hence addition of sucrose was done after 9th daysof progressive leaching.

Organic acids are chelators which can form complexes withmetals. Such complexation is dependent on the relative concen-trations of anions and metals in solution, the pH and the stabilityconstant of the various complexes (Denevre et al., 1996). Mc Kenzieet al. (1987) reported that solubilisation of nickel, cobalt and ironfrom laterites by means of organic chelating acids at a low pH.Productions of organics acids in the culture medium lowers the pHthereby induces the dissolution of metals. This fact also holds truewith our findings where A. niger produces maximum oxalic acidsand biomass showing maximum extraction of nickel and cobalt.

3.8. Reaction kinetics

Themineralogical analysis reveals that the nickel was associatedwith the goethite iron matrix of COB. Nickel solubilizes due to theattack of fungal bio acids from the iron matrix of COB. Kinetics ofthe bioleaching process has been studied to support the mecha-nism of the reaction. Various kinetic models such as chemical,diffusion and mixed control were tested.

If the rate of metal extraction is controlled by the chemicaldissolution of the ore particles by bio-acids generated during fungalbioleaching, then the following rate equation is generally applicable,by assuming spherical shape of the ore particle (Habasi, 1969).

Fig. 9. (a & b): Graph showing kinetics of Nickel extraction from pre-treated COB inshake flask (a) Application of Eq.(2) for initial rate of extraction (b) Application ofEq.(3), for overall rate of extraction.

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1� ð1� aÞ1=3 ¼ k1t (5)

However, if the nickel diffusion is very fast the kinetics ofleaching usually obey three dimensional diffusion equation of thefollowing type.

1� 2=3a� ð1� aÞ2=3 ¼ k2t (6)

Where a is leaching fraction of nickel at time t, k1 and k2 arereaction constants.

Fig. 10. (a & b): Application of Eq.(2) for cobalt extraction from pre-treated COB in (a)shake flask (b) Bioreactor.

Interpretation of the data obtained from bioreactor leachingshows that there was tremendous rise in the leaching percentageup to 9th day due to the consumption of sucrose by the fungalstrain. Detail explanation was given in Section 3.6. Therefore,initially the kinetics of nickel extraction was believed to becontrolled by chemical dissolution model as shown in (Fig. 8a) butsoon after addition of sucrose to the leaching medium there wasrise in the leaching percentage which later on fits to the threedimensional shrinking core model as depicted from (Fig. 8b).Therefore, considering the above facts it can be concluded that thereaction kinetics for nickel extraction follows a mixed model.Similar findings were also observed in shake flask (Fig. 9a & b).Whereas this does not holds true for cobalt in both shake flask andbioreactor leaching, where the reaction kinetics strictly followsa chemical control model. This might be due to the fact that, ascobalt is associated with minor manganese phase. Due to pre-treatment, it comes to a thermodynamically stable configurationwhere manganese occurs as an independent phase bearing cobalt.Hence, it was more susceptible to fungal acids in compare to rawCOB (Fig. 10a & b).

4. Conclusion

Chemical analysis of raw and pre-treated COB reveals that0.99% of nickel and 0.03% of cobalt in raw and 1.02% of nickel and0.04% of cobalt contains in pre-treated COB respectively. The XRDanalysis of chromite overburden showed nickel present in absor-bed state within the goethite matrix whereas in pre-treated COBrevealed conversion of goethite to hematite due to thermal pre-treatment at 600 �C under normal atmospheric conditions takesplaces. This result in increased leach-ability might be due toincreased porosity, increased surface area and change in reactivityof the ore due to extensive dehydration and dehydroxylation ofvarious hydrated oxide minerals. Efficiency for production oforganic acid of A. nigerwas also tested. TEM analysis of the leachedsamples shows that during progressive leaching nickel dissoluteinto the solution leaving a porous space in the matrix. Nickeldissolutes from the core of the hematite matrix in the pre-treatedCOB by forming organo-metallic complex (Ni-oxalate/Ni-citrate).Kinetics of the nickel bioleaching was studied to support themechanism of the reaction. The reaction kinetics for both shakeflask and bioreactor follows amixed control model in case of nickelextraction. Comparing this with cobalt recovery it was found outthat in both the cases it follows the chemical control model ascobalt is associated with minor manganese phase which comes toa thermodynamically stable configuration due to pre-treatmentprocess resulting into an independent phase bearing cobalt.Hence, it was more susceptible to fungal acids in compare to rawCOB. A maximum 18% of nickel and 37.8% of cobalt was recoveredfrom pre-treated COB at 2.5% pulp-density with 10% (v/v) fungalinoculum at 30 �C for duration of 25 days in shake flask whileunder same mentioned conditions, 32.5% of nickel and 86% ofcobalt was recovered in bioreactor.

Acknowledgement

The authors acknowledge Orissa Mining Corporation forproviding the samples. One of the authors would like to thanks CSIRfor awarding Senior Research Fellowship.

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