Development of a Laboratory-Scale Leaching Plant for Metal ...
<ul><li><p>APPLIED AND ENVIRONMENTAL MICROBIOLOGY,0099-2240/98/$04.0010</p><p>Apr. 1998, p. 12371241 Vol. 64, No. 4</p><p>Copyright 1998, American Society for Microbiology</p><p>Development of a Laboratory-Scale Leaching Plant for MetalExtraction from Fly Ash by Thiobacillus Strains</p><p>CHRISTOPH BROMBACHER,1 REINHARD BACHOFEN,1 AND HELMUT BRANDL2*</p><p>Institute of Plant Biology, Department of Microbiology, University of Zurich, CH-8008 Zurich,1 andInstitute of Environmental Sciences, University of Zurich, CH-8057 Zurich,2 Switzerland</p><p>Received 23 September 1997/Accepted 5 January 1998</p><p>Semicontinuous biohydrometallurgical processing of fly ash from municipal waste incineration was per-formed in a laboratory-scale leaching plant (LSLP) by using a mixed culture of Thiobacillus thiooxidans andThiobacillus ferrooxidans. The LSLP consisted of three serially connected reaction vessels, reservoirs for a flyash suspension and a bacterial stock culture, and a vacuum filter unit. The LSLP was operated with an ashconcentration of 50 g liter21, and the mean residence time was 6 days (2 days in each reaction vessel). Theleaching efficiencies (expressed as percentages of the amounts applied) obtained for the economically mostinteresting metal, Zn, were up to 81%, and the leaching efficiencies for Al were up to 52%. Highly toxic Cd wascompletely solubilized (100%), and the leaching efficiencies for Cu, Ni, and Cr were 89, 64, and 12%, respec-tively. The role of T. ferrooxidans in metal mobilization was examined in a series of shake flask experiments.The release of copper present in the fly ash as chalcocite (Cu2S) or cuprite (Cu2O) was dependent on themetabolic activity of T. ferrooxidans, whereas other metals, such as Al, Cd, Cr, Ni, and Zn, were solubilized bybiotically formed sulfuric acid. Chemical leaching with 5 N H2SO4 resulted in significantly increased solubi-lization only for Zn. The LSLP developed in this study is a promising first step toward a pilot plant with a highcapacity to detoxify fly ash for reuse for construction purposes and economical recovery of valuable metals.</p><p>Biohydrometallurgy, an interdisciplinary field involvinggeomicrobiology, microbial ecology, microbial biochemistry,and hydrometallurgy (23), is a novel promising technology forrecovering valuable metals from industrial waste materials(e.g., bottom and fly ash, galvanic sludge, and filter dust) andfor detoxifying these materials for environmentally safe depo-sition. Biohydrometallurgical processing of solid waste allowsrecycling of metals, similar to natural biogeochemical metalcycles, and diminishes the demand for resources, such as ores,energy, and landfill space. Fly ash from municipal waste incin-eration (MWI) is a concentrate containing a wide variety oftoxic heavy metals (e.g., Cd, Cr, Cu, and Ni). The zinc concen-trations in fly ash (3%, wt/wt) can be similar to the concentra-tions in ores subjected to conventional mining (16), whichmakes MWI fly ash a suitable candidate for economical zincrecovery. The low acute and chronic toxicity of fly or bottomash for a variety of microorganisms (8) and the low mutageniceffect (17) of this material have been demonstrated previously.However, the deposition of materials containing heavy metalsresults in a severe risk that spontaneous metal leaching mayoccur due to natural weathering processes and uncontrolledbacterial activities (18, 21, 23). Agenda 21 adopted at the 1992Earth Summit in Rio de Janeiro, Brazil, established that thereis a strong requirement to support sustainable development,including ecological treatment of wastes and safe disposal ofwastes. Biological metal leaching of fly ash is a step in thisdirection.</p><p>Biohydrometallurgy is a technology that is cleaner and con-sumes less energy than technologies used in the pyro- andhydrometallurgical industries. The latter technologies are well-established, and many of them are patented, whereas patentsfor biohydrometallurgical processing of industrial wastes are</p><p>rarely published (7). The first effort to develop biohydromet-allurgical treatment of industrial waste was made 20 years ago,and greater efforts are necessary now. This is an importantsubject of research and should result in a wide range of inves-tigations and applications in the future. However, the previousdata on biohydrometallurgical treatment of fly ash or otherindustrial waste obtained with bacteria or fungi included resi-dence times for leaching of up to 50 days (46, 11, 25). Most ofthese experiments were performed on a small scale with lowamounts of heavy-metal-containing material.</p><p>In this paper, a semicontinuous laboratory-scale leachingplant (LSLP) is described; this LSLP achieved high leachingefficiencies, which resulted in an elevated load of elements inthe effluent. Treatment times were found to be less than treat-ment times obtained with batch extraction procedures. A mix-ture of Thiobacillus thiooxidans producing sulfuric acid andThiobacillus ferrooxidans oxidizing reduced metal compounds(19) was used to perform the leaching experiments. The resultswere compared to chemical (abiotic) leaching efficiencies. Inaddition, we investigated whether T. ferrooxidans was neededfor leaching of fly ash. Metals can be biotically released fromfly ash by mechanisms such as direct enzymatic reduction,indirect action resulting from extracellular metabolic products,or acid formation (nonenzymatic dissolution), as previouslyshown in an anaerobic system (10). It was possible to differ-entiate among these release mechanisms in an oxic acidic flyash-Thiobacillus system.</p><p>MATERIALS AND METHODS</p><p>Bacterial strains, medium, and culture conditions. T. thiooxidans DSM622and T. ferrooxidans DSM2391 were cultivated in a medium containing (per liter)0.1 g of K2HPO4, 0.25 g of MgSO4 z 7H2O, 2.0 g of (NH4)2SO4, 0.1 g of KCl, and8.0 g of FeSO4 z 7H2O. Elemental sulfur (1%, wt/vol) was added, and the pH wasadjusted with sulfuric acid to 2.5 to 2.7. The mixed culture was grown undernonsterile conditions either in 250-ml baffled Erlenmeyer flasks on a rotaryshaker (140 rpm) or in an aerated and stirred 1,000-ml beaker. Growth wasmonitored by monitoring the pH (with a Hamilton single-pore electrode), thecell counts (with a Neubauer counting chamber), and the Fe(II) concentration(12).</p><p>* Corresponding author. Mailing address: Institute of Environmen-tal Sciences, University of Zurich, Winterthurerstrasse 190, CH-8057Zurich, Switzerland. Phone: 41 1 635 61 25. Fax: 41 1 635 57 11. E-mail:HBRANDL@UWINST.UNIZH.CH.</p><p>1237</p><p> on April 8, 2018 by guest</p><p>http://aem.asm</p><p>.org/D</p><p>ownloaded from</p><p>http://aem.asm.org/</p></li><li><p>Samples and sample preparation. A 500-kg portion of fly ash retained byelectric filters at the MWI plant in Hinwil, Switzerland, was collected by workersat Sulzer Chemtech (Winterthur, Switzerland) at different times on one day andwas homogenized in a cement mixer to obtain representative homogeneoussamples. The ash was washed with water to remove water-soluble compoundsand dried on a vacuum filter. For experiments the ash was ground and dried at80C for 48 h. The concentrations of selected elements are listed in Table 1. Thevalue for loss of combustion at 950C represents the inorganic carbon content.</p><p>Semicontinuous LSLP. The LSLP consisted of three serially connected reac-tion vessels (designated RV-A, RV-B, and RV-C), each having a volume of 1dm3 (Fig. 1). Pulp from the fly ash storage solution (100 g liter21) and thebacterial stock culture (109 cells/ml) were mixed in equal amounts and fed every12 h semicontinuously into the first reaction vessel (RV-A) with a peristalticpump at an overall dilution rate of 0.021 h21 (0.5 day21). RV-A and RV-B wereconnected by an overflow connector. The pulp was pumped from RV-B to RV-Cwith a peristaltic pump at the same rate to flush settled fly ash particles. This</p><p>resulted in a pulp concentration in the LSLP of 50 g liter21 and a mean residencetime of 6 days (2 days in each reaction vessel). The bacterial stock culture and thethree reaction vessels were aerated at a rate of about 2 volumes of air per volumeof reactor per min. After RV-C, the pulp was transported by gravity flow into a2-liter vacuum glass filter unit, and the particle-free, metal-rich solution wascollected in a 5-liter collecting vessel.</p><p>Determination of mobilization mechanisms. Forty milliliters of an 8% (wt/vol)fly ash suspension (acidified with sulfuric acid to pH 5.4) was diluted with 40 mlof a Thiobacillus culture (109 cells/ml) after different treatments (see below) andincubated for 8 days on a rotary shaker (150 rpm) at room temperature (23 to24C). All samples were incubated in triplicate. Several mechanisms of metalmobilization were distinguished, as described below.</p><p>The direct enzymatic effect on the release of metals was determined by dilutingthe ash suspension with bacterial stock cultures (pH 1.1). The cells were in directcontact with the fly ash. Growth of T. ferrooxidans might have been stimulated byincreased energy available from oxidation of reduced solid particles.</p><p>Leaching with cell-free spent medium revealed that indirect solubilization byextracellular metabolic products occurred. The stock culture was centrifuged at23,700 3 g, and the supernatant was filtered through a 0.22-mm-pore-size Teflonfilter to obtain cell-free spent medium. The cell-free spent medium was checkedfor viable cells by incubating a 5-ml sample in 80 ml of fresh medium.</p><p>Cell-free spent medium (see above) was autoclaved (12 min, 121C) to obtaina sterile leaching solution without enzymatic activities to evaluate the leachingability of the acid formed. The solution was checked for precipitates and forchanges in redox state after the heat treatment.</p><p>Forty milliliters of fresh uninoculated medium was added to the fly ash sus-pension and used as a control.</p><p>Elements such as Cd or Zn might have been chemically mobilized duringpreparation of the ash suspension due to the acidification to pH 5.4.</p><p>Chemical leaching with sulfuric acid. Eighty milliliters of the fly ash suspen-sion was leached at a concentration of 5% (wt/vol) with a maximum of 11 ml of5 N sulfuric acid (final pH, pH 2) at the following pH values: at an initial pH of10, at pH 8 (corresponding to the carbonate buffer pH), at pH 4 (correspondingto the potassium-aluminum buffer pH) (2), and at pH 2 (a possible end point ofa leaching experiment). The concentrations of the solubilized metals and the acidconsumption were measured. The pH at each value was controlled with a pH-Stat (Metrom Impulsomat model 614). Fly ash was suspended in distilled waterand stirred at a constant pH for 24 h. A new suspension was prepared for eachpH step.</p><p>Analytical procedures. Metal analyses were performed by using inductivelycoupled plasma atomic absorption spectroscopy (ICP-AES; Spectro AnalyticalInstruments, Kleve, Germany) and standard addition methods at the followingwavelengths: Al, 396.2 nm; Cd, 228.8 nm; Cr, 267.7 nm; Cu, 324.8 nm; Fe, 261.2nm; Mn, 294.9 nm; Ni, 352.5 nm; and Zn, 206.2 nm. Prior to the inductivelycoupled plasma analysis, the samples were centrifuged at 23,700 3 g for 15 min,acidified with 5 drops of concentrated HNO3 per 30 ml of aqueous solution,passed through a glass fiber filter (Whatman type GF/C) to guarantee particle-free suspensions, and stored at 4C.</p><p>RESULTS AND DISCUSSION</p><p>LSLP. A semicontinuous three-stage leaching plant for ex-traction of heavy metals from MWI fly ash was developed.Biohydrometallurgical processing of fly ash from MWI poses,especially at high pulp densities, severe problems due to thehigh content of toxic metals in the fly ash and the saline andstrongly alkaline environment. It is necessary to obtain reducedtreatment times for high pulp concentrations without addi-tional acidification; this is important for reducing the capitaland maintenance costs of a pilot plant.</p><p>RV-A of the LSLP (Fig. 1) was filled with 500 ml of abacterial culture (pH 1.5) and 250 ml of a fly ash suspension(10%, wt/vol) for the first adaptation phase. After 36 h, an-other 250-ml aliquot of the suspension was added to obtain thefinal 5% (wt/vol) solution. When the pH in RV-A droppedbelow 2, the LSLP was started. After 48 h (corresponding tofour discontinuous feeding cycles) the solution was pumped toRV-C. After 48 h, the microorganisms produced sufficientamounts of sulfuric acid in each reaction vessel to maintainsteady-state conditions despite the alkaline pH of the fly ashpulp (pH 9 during the experiment). A distinct pH cascadeoccurred from one reaction vessel to the following reactionvessel. The pH fluctuated during the steady state depending onthe fly ash present; in RV-A the pH fluctuated between 3.7 and4, in RV-B the pH fluctuated between 2.7 and 3.2, and in RV-C</p><p>FIG. 1. Schematic diagram of the LSLP. A, B, and C, serially connectedRV-A, RV-B, and RV-C, respectively; 1, fly ash reservoir; 2, bacterial stockculture; 3, peristaltic pump; 4, filter unit; 5, collecting vessel; 6, to vacuum pump;solid lines, liquid flow; dashed lines, air flow.</p><p>TABLE 1. Concentrations of selected elements in fly ash from theMWI plant in Hinwil, Switzerlanda</p><p>Element Detectionmethod(s)bConcn in flyash (g kg21)</p><p>SE(%)</p><p>Al A, B, C 70 5.5Ca A, B, C 132 5.5Cd B 0.49 15.0Cl B 4.7 15.0Cr A, B 0.7 7.9Cu B 1.1 15.0F B 8.0 15.0Fe A, B, C 28 5.5Hg B ,0.01 15.0K A, B, C 12 5.5Mg A, B, C 14 5.5Mn A, B, C 0.77 5.5Na A, B, C 11 5.5Ni A, B 0.14 7.9Pb B, C 8.9 11.1S B, D 30 7.9Si A, B 100 7.9Sn B 9.3 15.0Ti A, B 12 7.9V B 0.13 15.0Zn B, C 31 7.9Loss of combustion (1 h, 950C) 126 NDc</p><p>a Data reproduced from reference 27 with permission. The measured concen-trations allowed us to determine approximate leaching efficiencies.</p><p>b A, X ray fluorescence with a glass specimen (standard error, 65%); B, X rayfluorescence with a compacted powder specimen (standard error, 615%); C,inductively coupled plasma atomic absorption spectroscopy (standard error,65%); D, infrared detection after oxidation at 2,000C (standard error, 65%).</p><p>c ND, not determined.</p><p>1238 BROMBACHER ET AL. APPL. ENVIRON. MICROBIOL.</p><p> on April 8, 2018 by guest</p><p>http://aem.asm</p><p>.org/D</p><p>ownloaded from</p><p>http://aem.asm.org/</p></li><li><p>the pH fluctuated between 1.2 and 1.5. Before the bacterialstock was added to RV-A, growth parameters [pH, Fe(II)concentration, cell number] were monitored (Fig. 2). The me-dium was fully replenished with fresh medium every 72 h (cor-responding to an overall dilution rate of 0.01 liter h21) to avoidaging of the bacterial stock culture.</p><p>Samples used for metal analysis were removed after 168 hfrom each reaction vessel, from the collection vessel, and (ascontrols) from the fly ash storage vessel and the bacterial stockculture. For all elements, the concentrations of soluble metalsincreased continuously with increasing mean residence time inthe LSLP (Fig. 3). In RV-C the following amounts of metalswere solubilized (per kilogram of fly ash): Al, 37 g; Zn, 25 g;Fe, 3.1 g; Cu, 0.98 g; Mn, 0.53 g; Cd, 0.49 g; Ni, 0.09 g; and Cr,0.08 g. Ferrous iron added to the bacterial stock culture as anelectron...</p></li></ul>