Coal Desulfurization with

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Coal Desulfurization with Acidithiobacillus ferrivorans, from Balya Acidic Mine Drainage Pinar Aytar,*,† Catherine M. Kay,‡ Mehmet Burçin Mutlu,§ and Ahmet Ç abuk∥ †

Graduate School of Natural and Applied Sciences, and ∥Department of Biology, Faculty of Arts and Science, Eskisehir Osmangazi University, 26480 Eskisehir, Turkey ‡ Environmental Microbiology, Bangor Acidophile Research Team, Bangor University, Gwynedd LL57 2UW, United Kingdom § Department of Biology, Faculty of Science, Anadolu University, Eskisehir 26470, Turkey ABSTRACT: The biodesulfurization capability of a strain having sulfur and iron metabolism isolated from acidic mine drainage of Balya (Balikesir, Turkey) was studied. Molecular identification of the 16S rRNA gene showed that this bacterium was a strain of Acidithiobacillus ferrivorans. Desulfurization optimization experiments were performed by Taguchi’s method. Statistical experimental arrangement L16 (45) was prepared to determine optimum sulfur removal. The optimum conditions for these parameters were found to be pH of 2.5, inoculum amount of 2%, pulp density of 1%, particle size of −500 + 250 μm, and incubation time of 14 days. A value of “Prob > F” less than 0.0500 indicates that model terms are significant. The obtained yields of total sulfur removal were approximately 33%. According to variance analysis, it was seen that all parameters were effective in removal of total sulfur. Scanning electron microscopy and Fourier transform infrared spectroscopy analyses also indicated a modification of the coal surface after biodesulfurization. The redox potential was measured as 818 mV (7 days) and 788 mV (14 days) during the biodesulfurization experiment by the Pt−Ag/AgCl system of cyclic voltammetry, which suggested that the Fe3+/Fe2+ redox pair could be thermodynamically competitive with the O2/H2O couple as the electron acceptor during bacterial sulfur oxidation, demonstrating that S0 oxidation was coupled with Fe3+ reduction. Thermogravimetry, differential thermal analysis, and differential thermogravimetry curves for untreated and biotreated coal showed the differences in combustion profiles, possibly relating to structural alterations derived from biotreatments. respire ferric iron with elemental sulfur anaerobically.10 A. ferrivorans SS3, isolated from Siberia is able to oxidize ferrous iron and tetrathionate as well as catalyze the leaching of pyrite/ arsenopyrite and chalcopyrite at +4 °C considerably faster than A. ferrooxidans.12,13 In this study, sulfur of coal from Turkey was removed using a pure culture of A. ferrivorans isolated from Balya acidic mine drainage. The first target of this work is to optimize the desulfurization conditions of A. ferrivorans using Taguchi’s methodology. The second objective was to determine mineralogical transformations in coal occurring during the biodesulfurization process. To establish physical, chemical, and thermal behavior changes in the coal, characterization techniques, such as Fourier transform infrared (FTIR) spectroscopy, thermogravimetric analysis (TGA)/differential thermal analysis (DTA), and scanning electron microscopy (SEM) analyses, and chemical measurements, such as pH, Eh, and concentration of iron in solution, were carried out.

1. INTRODUCTION Acid mine drainage (AMD) containing an abundance of FeII and FeIII is produced when sulfide minerals are exposed to oxidizing conditions and has lower acidic pH values and high specific conductivity. The source of acidity is caused by the oxidation of sulfide compounds, especially pyrite. Although this process occurs naturally, the mining process can also promote AMD generation. Moreover, naturally occurring bacteria can accelerate AMD production by assisting in the breakdown of sulfide minerals.1 Some acidophilic microorganisms found in AMD waters play a role as natural catalysts for several of the oxidation reactions involved in AMD formation. Earlier studies of the microbiology of AMD systems paid attention almost completely on the prokaryotic populations, such as bacteria and archaea, because of biotechnological reasons, such as bioleaching, metal bioaccumulation, and biodesulfurization.2 Biodesulfurization is an eco-friendly approach for fossil fuel because of its advantages, such as low energy consumption and production of non-hazardous byproducts.3,4 Although certain microorganisms or their enzymes are found to remove sulfur compounds,5 the most studied microorganism for metabolizing these compounds is the chemoautotroph Acidithiobacillus ferrooxidans.6−9 A. ferrivorans growing in the range of 5−37 °C, is the newest member of the Acidithiobacillus genus.10 It is a member of phylogenetic group III of the iron-oxidizing Acidithiobacillus genus, whereas A. ferrooxidans is group I.11 Furthermore, it can oxidize iron and reduce sulfur compounds aerobically and © XXXX American Chemical Society

2. EXPERIMENTAL SECTION 2.1. Coal Samples. Coal samples from Mihaliccik open mine were used for desulfurization experiments. Proximate, ultimate, and petrographic analyses of the coal were carried out with Leco TGA500 for calorimetric analysis and Leco TGA700 for ash and volatile matters. Total sulfur analysis was performed using an Eltra CSReceived: March 2, 2013 Revised: May 29, 2013

A

dx.doi.org/10.1021/ef400360t | Energy Fuels XXXX, XXX, XXX−XXX

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(medium pH, coal particle size, inoculum amount, pulp density, and incubation time) at four levels with an orthogonal array layout of L16 (45). All statistical experimental results were analyzed using Minitab 16 for Windows (Minitab, Inc.). Working conditions for all of the samples were a 250 mL flask with a medium volume of 100 mL. The agitation rate was kept stable at 125 rpm. For each optimization stage, at the end of the incubation time, the coal samples were separated by filtration and then agitated with hot 15% HCl for removing jarosite salts, which are unwanted compounds, and washed with distilled water. The coal samples were dried at 45 °C and analyzed for total sulfur content. The extent of desulfurization was followed by an analysis of the total sulfur in the coal samples after microbial treatments, and the calculated percentage was found as the difference in the total sulfur content of coal before and after treatment. 2.5. Analytical Measurements. To observe changes in chemical bonds, FTIR spectroscopy was used. The FTIR spectra of the coal before and after biodesulfurization were recorded using a Bruker Tensor 27 spectrophotometer in the region of 400−4000 cm−1. FTIR spectra were measured on KBr pellets prepared by pressing mixtures of 1 mg of dry powdered sample and 100 mg of spectrometry-grade KBr under vacuum. SEM analysis was carried out to observe possible changes on the particle surface after desulfurization with a JEOL 5600 LV SEM instrument. Most of the microorganisms having an ability of desulfurization are known to oxidize also ferrous iron. To measure changes in the ferrous iron concentration, analyses using the ferrozine assay and cyclic voltammetry (CV) were used. According to the ferrozine colorimetric technique suggested by Lovley and Phillips, iron oxidation was monitored in culture flasks after desulfurization;22 total iron was quantified by the same method after reduction with hydroxylamine. The difference between ferrous iron and total iron was taken as ferric iron. For CV experiments, the electrochemical measurements were carried out using a Gamry 3000 potentiostat/galvanostat/ZRA system (Warminster, PA) and obtained data were analyzed using Gamry CMS-300 (version 5.50b) framework/analysis software. All experiments were carried out at 25 °C. CV measurements of untreated and treated desulfurization medium were performed in the potential region between −0.30 and +1.10 V versus Ag/AgCl at 50 mV/s scan rate over 15 cycles. To understand the thermal behavior of coal before and after biodesulfurization, a Perkin-Elmer Diamond TG/DTA thermal analyzer was used to record simultaneous thermogravimetry (TG), differential thermogravimetry (DTG), and DTA curves. The samples (20−25 mg, −100 μm of particle size) are heated to 800 °C at a rate of 10 °C/min, individually in the atmosphere of a dry air flow rate of 50 cm/min using platinum crucibles.

530 sulfur analyzer with an infrared absorption detection procedure. The sulfur emission value (EV), which is grams of sulfur per megajoule (g of S/MJ), was calculated by dividing the total sulfur content (wt %) by the calorific value (MJ).8 Also, the sulfur forms, such as pyritic, sulfatic, and organic, were determined by following ASTM 2492 at Standard Methods, and the elements, such as C, H, and N, were determined through the method suggested by ASTM 5291.14 The maceral composition of this coal sample was determined by point counting the polished briquettes produced from the reflectance measurements. The terminology used for maceral identification is adopted from Stach et al.15 After coal was dried at 105 °C, incinerated at 550 °C, and digested according to TS ISO 11466, major elements of this sample were analyzed using inductively coupled plasma−optical emission spectrometry (Perkin Elmer 400). For minor element analysis, XRF equipment (Thermo-ARL) was used. Table 1 shows characteristics of the coal.

Table 1. Characteristics of the Coal calorific value (cal/g) moisture content (%) ash (%) volatile matter (%) fixed carbon (%) Na2O (%) MgO (%) Al2O3 (%) SiO2 (%) K2O (%) P2O (%) CaO (%) TiO2 (%) SO3 (%) Fe2O3 (%)

2152 3.85 54.70 24.70 16.75 8.7 2.5 14.0 43.1 0.6 0.2 6.9 0.9 10.1 7.2

MnO (%) As (ppm) Ba (ppm) Cr (ppm) Cu (ppm) Mo (ppm) Pb (ppm) Sr (ppm) Rb (ppm) Th (ppm) U (ppm) V (ppm) Y (ppm) Zn (ppm)

pulp density > coal particle size > pH > inoculum amount. The Δ values of pH, coal particle size, inoculum amount, pulp density, and incubation time are 8.947, 9.148, 7.547, 9.224, and 14.499 (Table 3). Values of “Prob > F” less than 0.0500 indicate that model terms are significant. In this case, all factors are significant model terms. Table 4 indicates variance analysis according to the general linear model.

species is a psycrotolerant facultative anaerob, obligately chemolithotrophic and disazotrophic.10 3.2. Optimization of Sulfur Removal Using Taguchi’s Methodology. As shown in Table 2, the effects of five factors on desulfurization, pH (A), coal particle size (B), inoculum amount (C), pulp density (D), and incubation time (E), were studied. Taguchi’s L16 orthogonal array having four levels for each of these variables was performed, and parameters were selected using information obtained from our previous studies.4 For the Taguchi design and analysis, the Minitab Statistical Software package was used. Desulfurization experiments were carried out with the isolate performing the 16 trials consisting of triplicate experiments, and percent average desulfurization values were given in Table 2. For determination of the best process parameters, the larger the better case was used (Figure 1). In the experiments for total sulfur removal, pH of 2.5, C


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a high huminite content (75%), 5% liptinite, 3−4% inertinite, 0.0524% organic matter, 10% clay, quartzous, and calcite minerals, and 6−8% framboidal pyrite and iron hydroxide minerals; therefore, it has sub-bituminous characteristics. Table 5 shows the analysis of coal samples before and after the biodesulfurization process by A. ferrivorans and the

Table 3. Response Table for Means factors

levels

A (pH)

B (coal particle size)

1 2 3 4 Δ rank

11.307 13.596 13.072 4.649 8.947 4

6.094 8.943 12.345 15.242 9.148 3

C (inoculum amount)

D (pulp density)

E (incubation time)

6.184 13.732 9.613 13.095 7.547 5

15.187 10.638 6.663 9.436 9.224 2

7.760 19.991 9.382 5.491 14.49 1

Table 5. Comparison of Untreated and Treated Coal in Terms of Coal Characteristics before biodesulfurization (untreated coal)

after biodesulfurization (treated coal)

3.42 2152

2.29 2056

380

286

2.37 0.72 0.33 3.85 54.70 24.70 16.75 26.95 2.13 1.27

1.42 0.65 0.22 3.53 50.54 21.44 24.49 21.5 1.79 1.05

total sulfur (wt %) calorific value (cal/g) sulfur emission value (EV) pyritic sulfur (%) organic sulfur (%) sulfate (%) humidity ash (%) volatile matter (%) fixed carbon (%) carbon (%) hydrogen (%) nitrogen (%)

The interactive effects of the parameters were not taken into account in the theoretical analysis because our preliminary tests showed that they could be neglected. A verification experiment is a powerful tool for detecting the presence of interactions among the control parameters. To validate the optimum condition, the suggested optimum solution was performed again. The measured result of confirmation experiments, such as 33.04%, was found to be reasonably close to the predicted result (35.82%) and fell inside of the prediction interval, which confirmed the validity and adequacy of the predicted models. Several studies relating to biodesulfurization by Acidithiobacillus genus are available in the literature. However, A. ferrivorans was first used for sulfur removal from coal in this study. In another study with Acidithiobacillus caldus, the results indicated that total desulfurization was about 19%. After processing for 40 days, the cells attached to the surface of pyrite and there was clear corrosion on the mineral surface.26 Malik and co-workers applied processes consisting of three steps through A. ferrioxidans; at the end of the last step, 51% of pyrite removal for 10 days was obtained.27 In another study, treatment of a type of lignite by Pseudomonas putida B2 attained 44% of total sulfur removal, while total sulfur removal of a subbituminous coal by A. ferrooxidans F3 attained 14%.28 Acharya et al. studied different coal biodesulfurization by A. ferrooxidans; the conditions studied were optimized for the maximum removal of sulfur (91.81 wt % for Rajasthan lignite, 63.17 wt % for Polish bituminous coal, and only 9.41 wt % for Assam coal). The cause of dissimilarities of the removal rate obtained might be differences of the rank of coal studied.7 In this study, in addition, a significant desulfurization performance was exhibited by A. ferrivorans; because of having advantageous characteristics, including a psycrotolerant feature, it has more potential for biotechnological applications, such as biodesulfurization.10 3.3. Changes in Coal before and after Biodesulfurization. 3.3.1. Characterization of the Coal Sample and Changes of Coal Quality. Petrographic analysis of the coal sample used in this study indicated that it consists of groups of

characteristics of coal; the working conditions of the coal were pH of 2.5, inoculum amount of 2%, pulp density of 1%, particle size of −500 + 250 μm, and incubation time of 14 days, which were optimal conditions found as a result of the Taguchi statistical method. The result of the confirmation experiment was 33.04% of total sulfur removal; besides, the pyritic sulfur for the same coal sample was reduced from 2.37 to 1.42%, and the pyritic desulfurization was 40.08%. However, small alterations for organic and sulfatic sulfur were observed. Other characteristics of the coal sample are represented in Table 5. While the calorific value did not significantly change, the emission value, a factor of the environmental effect, also decreased. 3.3.2. FTIR Analysis. FTIR spectra for raw coal and the biodesulfurized sample are shown in Figure 2. In addition to mineralogical information, although this technique can be used to recognize organic compounds in the coal structure, the major difficulty when the raw coal sample is analyzed is that a superim position of organic and inorganic bands is common.29 Alterations of chemical structures in coal before and after biodesulfurization were presented in Figure 2. The band shifts that occurred in several regions may suggest some information about sulfur removal of coal.

Table 4. Variance Analysisa

a

parameter

freedom rate

seq SS

adj SS

adj MS

F factor

p value

A B C D E error total

3 3 3 3 3 32 47

611.81 571.52 437.94 537.54 1485.87 136.96 3781.64

611.81 571.52 437.94 537.54 1485.87 136.96

203.94 190.51 145.98 179.18 495.29 4.28

47.65 44.51 34.11 41.87 115.72

0.000 0.000 0.000 0.000 0.000

S, 2.068 80; R2, 96.38%; and R2(adj), 94.68%. D


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Figure 2. FTIR spectra (a) before and (b) after desulfurization.

At the end of the 10th day, Fe2+ amount reduction was 88%. When the 14th day was reached, this value increased up to 99.73%. From the results, it is clear that the pyrite in the coal had been used throughout the biodesulfurization process. The coal used in this study had a content of pyritic sulfur (FeS2) of approximately 69%; therefore, the iron removal rate as well as sulfur would be greater. On the basis of these results, it is remarkable that the cells removed the majority of pyrite from the coal sample. Pyritic sites are more susceptible to bacterial oxidation.32 Indeed, Fe2+ measurement and CV experiments were performed because there were various Fe−S compounds in coal. In this context, there is no stoichiometric correlation between sulfur and iron removal because iron is not only referring to pyrite but also to other Fe compounds not even containing sulfur. During biodesulfurization, the redox potential was measured as 818 and 788 mV at the end of the 7th and 14th days, respectively (Figure 4). It suggested that the Fe3+/Fe2+ redox pair could be thermodynamically competitive with the O2/H2O couple as an electron acceptor during bacterial sulfur oxidation; therefore, this demonstrated that S0 oxidation was coupled with

One of changes that occurred in the coal structure was an increase of intensity at 1035 cm−1 of the peak and shifting a band from 775 to 771 cm−1 after desulfurization, these band changes have been suggested as S−O stretching. Furthermore, the disappearance of bands at 1500 and 1703 cm−1 and only the appearance of bands at 1686 and 626 cm−1 may be assigned to stretching and an intensity change of C−O, S−O, N−O, and C−N bonds possibly found in coal. 3.3.3. SEM Analysis. The SEM images of untreated and biotreated coal particles were shown in panels a and b of Figure 3. In Figure 3a, the images indicate that the pyrite surface was smooth before desulfurization; however, it became rough with corrosive morphology (Figure 3b) after the desulfurization process for 14 days, especially larger holes where cells clearly attached. The attacks on the mineral surface were reported to occur via diffusion, convection, and chemotaxis.30 According to the study by Rohwerder et al., A. ferrooxidans preferentially attached to sites with visible surface imperfection.31 Similar results were reported in the study by He and colleagues.26 3.3.4. Iron Oxidation. The alterations of the amount of Fe2+ and total iron during biodesulfurization were shown in Table 6. E


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Figure 3. SEM analyses of coal (a) before and (b) after biodesulfurization.

Fe3+ reduction.13 It was reported that Acidithiobacillus strain SS3 was capable of using three various electron donor/acceptor pairs, such as Fe at suboptimal temperatures, Fe2+/O2, S0/O2, and S0/Fe3+.13 3.3.5. TG/DTA Analysis. Sulfur removal may improve coal combustion parameters. Thermal analytical methods, such as TG and DTA were shown to be effective at analyzing coal combustion behaviors.33,34 Panels a and b of Figure 5 show TG, DTA, and DTG curves for untreated and biotreated coal, respectively. Thermal changes in biotreated coals are generally

Table 6. Changes of Total Iron and Fe2+ during Desulfurization 10th day

untreated coal treated coal

14th day

total Fe (mg/L)

Fe2+ (mg/L)

Fe3+ (mg/L)

total Fe (mg/L)

3399

1325

2074

2531

3439

158

3281

1933

Fe2+ (mg/L) 1227 3.2

Fe3+ (mg/L) 1304 1929

F


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Figure 4. Cyclic voltammograms of culture solution for 7 and 14 days at a scan rate of 50 mV/s.

assigned to oxidation by microorganisms.35 As shown in these panels, the difference of weight losses between untreated and biotreated coal may be attributed to gases as a result of the biological process. The combustion profile of untreated and biotreated coal shows a single peak in the DTG curve temperatures at about 450 and 430 °C, respectively. The difference of weight loss between these DTG curves might also arise from other coal characteristics, such as ash content. The change in the reactivity parameter and thermograms of coal suggests a significant role for the petrographic components.36 A

similar difference of the temperature for decomposition of untreated and biotreated coal samples was reported in the study by Marinov et al.35 According to the results, the DTG curves of all coal samples demonstrated the major peaks of weight loss in the range of 350−400 °C. There was a general trend that the peak maximum of all biotreated samples is shifted to lower temperatures.35 DTA profiles visualize the heat flow with elevated temperatures. The first part of DTA curves may correspond to an endothermic process because of water evaporation. However, G


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Figure 5. TG/DTA and DTG profiles (a) before and (b) after biodesulfurization.

This study is the first report of the biodesulfurization of coal by A. ferrivorans under the depicted conditions. The experimental results from the optimization studies show that A. ferrivorans isolated from Balya AMD can be effectively used as a promising agent for the coal desulfurization process. A. ferrivorans could be applied as a desulfurization agent for a larger scale.

the main peak may be assigned to exothermic reactions (oxidation).35

4. CONCLUSION The effects of particle size, pulp density, initial pH, incubation time, and inoculum amount on the total sulfur removal of coal by a bacterial strain that isolated a Balya acidic mine drainage, A. ferrivorans PA8, were investigated at lab scale. In the experiments for total sulfur removal at the initial pH of 2.5, inoculum amount of 2%, pulp density of 1%, particle size of −500 + 250 μm, and incubation time of 14 days, the optimum conditions for the maximum reduction of total sulfur (33%) were found using Taguchi’s approach.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Notes

The authors declare no competing financial interest. H


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Microbe−Metal Interactions; Lovley, D. R., Ed.; American Society of Microbiology Press: Washington, D.C., 2000; pp 53−78. (25) Ghauri, M. A.; Okibe, N.; Johnson, D. B. Hydrometallurgy 2007, 85, 72−80. (26) He, H.; Hong, F.; Tao, X.; Li, L.; Ma, C.; Zhao, Y. Fuel Process. Technol. 2012, 101, 73−77. (27) Malik, A.; Dastidar, M. G.; Roychoudhury, P. K. J. Environ. Sci. Health, Part A: Toxic/Hazard. Subst. Environ. Eng. 2001, 36, 1113− 1128. (28) Marinov, S. P.; Stefanova, M.; Gonsalvesh, L.; Groudeva, V.; Gadjanov, P.; Carleer, R.; Yperman, J. Fuel Process. Technol. 2011, 92, 2328−2334. (29) Huggins, F. Int. J. Coal Geol. 2002, 50, 169−214. (30) Sand, W.; Gherke, T. Res. Microbiol. 2006, 15, 49−56. (31) Rohwerder, T.; Gehrke, T.; Kinzler, K.; Sand, W. Appl. Microbiol. Biotechnol. 2003, 63, 239−248. (32) Cardona, I. C.; Marquez, M. A. Fuel Process. Technol. 2009, 90, 1099−1106. (33) Cara, J.; Vargas, M.; Moran, A.; Gomez, E.; Martinez, O.; Frutos, F. J. G. Fuel 2006, 85, 1756−1762. (34) Kücu̧ ̈kbayrak, S. Thermochim. Acta 1993, 216, 119−129. (35) Marinov, S. P.; Gonsalvesh, L.; Stefanova, M.; Yperman, J.; Carleer, R.; Reggers, G.; Yürüm, Y.; Groudeva, V.; Gadjanov, P. Thermochim. Acta 2010, 497, 46−51. (36) Biswas, S.; Choudhury, N.; Sarkar, P.; Mukherjee, A.; Sahu, S. G.; Boral, P.; Choudhury, A. Fuel Process. Technol. 2006, 87, 191−199.

ACKNOWLEDGMENTS The study was supported by Eskisehir Osmangazi University Scientific Research Projects Committee (Project 201119018). This study is based partly on the Ph.D. thesis P. Aytar and she thanks TUBITAK (The Scientific and Technical Research Council of Turkey) BIDEB for a grant. We thank Dr. Barrie Johnson for isolation of the acidophile microorganism, Dr. Nimetullah Burnak and Yeliz Buruk for setup of experiments, and Dr. Derya Ö z Aksoy and Dr. Sabiha Koca for analyses of coal. Also, we express our gratitude to the Chemistry Department at Eskişehir Osmangazi University for TG/DTA, FTIR, and cyclic voltammetry analyses.

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