化工44-10表1̲4 2011/10/4 VOL. NO. C(DIC221) K 44 10 O C T O B E R 2 0 1 1 JOURNAL OF CHEMICAL ENGINEERING O F J A PA N [ VOL. 44, NO. 10, OCTOBER 2011 ] Special Issue for Sustainable Chemical Engineering Preface to the Special Issue for Sustainable Chemical Engineering Akihiro Yamasaki, Hideo Kameyama, Toshinori Kojima, Satoshi Nakai, Yuji Sakai and Mikiya Tanaka –––––––––––––––––– 677 Recent Advances in Extraction and Separation of Rare-Earth Metals Using Ionic Liquids (Journal Review) Yuzo Baba, Fukiko Kubota, Noriho Kamiya and Masahiro Goto ––––––––––––––––––––––––––––––––––––––––––––––––––– 679 Extraction Equilibria of Palladium(II) and Platinum(IV) with N,N-Di(2-ethylhexyl)aminomethylquinoline from Hydrochloric Acid Yoshinari Baba, Akemi Arima, Shintaro Kanemaru, Minako Iwakuma and Tatsuya Oshima ––––––––––––––––––––––––––– 686 Bacterial Cyanide Generation in the Presence of Metal Ions (Na, Mg2, Fe2, Pb2) and Gold Bioleaching from Waste PCBs Chi Dac Tran, Jae-Chun Lee, Banshi Dhar Pandey, Jinki Jeong, Kyoungkeun Yoo and Trung Hai Huynh ––––––––––––––– 692 The Society of Chemical Engineers, Japan Remediation of Heavy-Metal-Contaminated Soil with Chelating Agent and Treatment of Waste Solution Yoshiro Maki, Junji Shibata and Norihiro Murayama –––––––––––––––––––––––––––––––––––––––––––––––––––––––––––– 701 Phosphorus Recovery as Potassium–Magnesium–Phosphate from Effluent of Piggery Wastewater Using Zr(IV)-Loaded Saponified Orange Juice Residue Seiichirou Ohura, Biplob Kumar Biswas, Hiroyuki Harada, Mitsunori Kondo, Katsutoshi Inoue, Keisuke Ohto and Hidetaka Kawakita ––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––– 708 Nitrogen Transformations in Paddy Fields Treated with High Loads of Liquid Cattle Waste Sheng Zhou, Hiroshi Iino, Shohei Riya, Megumi Nishikawa, Yoichi Watanabe and Masaaki Hosomi ––––––––––––––––––– 713 A Metal-Monolithic Anodic Alumina-Supported Ag Catalyst for Selective Catalytic Reduction of NOx with Propene in the Presence of Hydrogen Yu Guo, Jian Chen and Hideo Kameyama ––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––– 720 Catalytic Oxidation of Volatile Organic Compounds over Co-Based Catalysts Supported on Charcoal from Thinned Wood Shimpei Kawada, Yu Guo, Lu Jia, Jian Chen, Mariko Kanehira, Tomohisa Kida, Koji Tsuboyama and Hideo Kameyama –––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––––– 729 Evaluation of Changes in SO2 Emissions and Economic Indicators Following the Reclamation of Alkali Soil in China Using By-Products of Flue Gas Desulfurization Yuji Sakai, Satoshi Nakano, Hirofumi Kito and Masayoshi Sadakata ––––––––––––––––––––––––––––––––––––––––––––––– 735 Desulfurization Performance of Sorbent Derived from Waste Concrete (SC) Atsushi Iizuka, Akihiro Yamasaki and Yukio Yanagisawa ––––––––––––––––––––––––––––––––––––––––––––––––––––––––– 746 Salt-Affected Soil Amelioration with Flue Gas Desulfurization By-Products and Waste Gypsum Board in Tianjin, China Yuji Sakai, Shunrong Ren, Chang Wang and Masayoshi Sadakata –––––––––––––––––––––––––––––––––––––––––––––––– 750 Effect of Coexisting FeO on Volatilization of PbO from CaO–SiO2–Al2O3–FeO Molten Slag in N2 Atmosphere Daisuke Shima, Hiroki Kageyama, Shohichi Osada, Mitsuhiro Kubota and Hitoki Matsuda ––––––––––––––––––––––––––– 757 JCEJAQ 44(10) 677-827(2011) ISSN 0021-9592 GIS-Based Estimation of Global Carbon Sequestration Potential Due to Forest Management Keigo Akimoto, Toshimasa Tomoda, Kiyotaka Tahara and Toshinori Kojima –––––––––––––––––––––––––––––––––––––––– 764 Influence of Gas Bubbling and Addition of Metal Oxide Particles on Ultrasonic Degradation of Methylene Blue Osamu Terakado, Ryo Sato and Masahiro Hirasawa –––––––––––––––––––––––––––––––––––––––––––––––––––––––––––– 774 This article appeared in the Journal of Chemical Engineering of Japan. The attached copy is provided to the author for non-commercial research, education use and sharing with colleagues. Other uses listed below are prohibited: - Reproduction, - Commercial use, - Posting to personal, institutional or third party websites. Research Paper Journal of Chemical Engineering of Japan, Vol. 44, No. 10, pp. 692–700, 2011 Bacterial Cyanide Generation in the Presence of Metal Ions (Naⴙ, Mg2ⴙ, Fe2ⴙ, Pb2ⴙ) and Gold Bioleaching from Waste PCBs Chi Dac TRAN1,2, Jae-Chun LEE2, Banshi Dhar PANDEY2,3, Jinki JEONG2, Kyoungkeun YOO4 and Trung Hai HUYNH5 1 University of Science and Technology (UST), Daejeon 305-350, Korea Korea Institute of Geoscience and Mineral Resources (KIGAM), Daejeon 305-350, Korea 3 National Metallurgical Laboratory (NML), CSIR, Jamshedpur, India 4 Department of Energy and Resources Engineering, Korea Maritime University, Busan, Korea 5 Institute for Environmental Science and Technology, Hanoi University of Science and Technology, Hanoi, Vietnam 2 Keywords: C. violaceum, Cyanide Generation, Bioleaching, Printed Circuit Boards, Gold As an alternative using cyanide chemicals for gold extraction, the application of a cyanogenic bacterium viz. Chromobacterium violaceum (C. violaceum) in YP medium has been investigated. The catalytic roles of metal ions such as Naⴙ, Mg2ⴙ, Fe2ⴙ, and Pb2ⴙ, as well as the effect of Na2HPO4 nutrient addition on the cyanide generation efficiency of the bacterium in this medium have been elucidated. While MgSO4 and FeSO4 added to the medium were equally effective for cyanide generation, improved efficiency was obtained in the presence of Na2HPO4 and Pb(NO3)2. In order to examine the effectiveness of C. violaceum cultured in YP medium for the generation of cyanide ions, the dissolution of gold and copper from waste mobile phone printed circuit boards (PCBs), a good source of gold and copper in alkaline conditions, was tested at 30°C, for various pH values and metal ion contents. Gold leaching was found to be 11% in 8 d at pH 11.0 in presence of 4.0 ⴛ 10ⴚ3 mol/L MgSO4, whereas; copper recovery was high (11.4%) at pH 10.0. Addition of 1.0 ⴛ 10ⴚ2 mol/L Na2HPO4 and 3.0 ⴛ 10ⴚ6 mol/L Pb(NO3)2 to the YP medium increased copper leaching to 30.3% and 38.1%, respectively, at pH 10.0 in 8 d. However, this effect was not observed for gold leaching. Introduction Cyanidation is widely used in gold metallurgy to recover gold from ores and concentrates, in spite of the fact that cyanides are highly toxic. The impact of cyanidation on the environment, especially on aquatic species and humans, is well known. In recent years, an alternate approach to the gold cyanidation process is being considered, in which the aim is to replace with a micro-organism, specifically cyanogenic bacteria that can produce cyanide ions and dissolve gold. So far, several types of cyanogenic bacteria have been identified and reported, which include Pseudomonas aeruginosa, P. fluorescens, Escherichia coli and Chromobacterium violaceum (C. violaceum) (Moss and Ryall, 1981; Creczynski-Pasa et al., 2004). Among these bacteria, C. violaceum has been found to be the most effective for the bio-dissolution of gold from different materials because of its cyanide-associReceived on October 1, 2010; accepted on July 7, 2011 Correspondence concerning this article should be addressed to J.-c. Lee (E-mail address: jclee@kigam.re.kr). 692 ated metabolic activities. In this study, C. violaceum has been cultured in YP medium containing glycine as one of the components. It is a mesophilic, motile, Gramnegative, and facultative anaerobe (Moss and Ryall, 1981). The importance attached to C. violaceum as a cyanide-generating bacterial species stems from its characteristic features of HCN synthase in comparison to other cyanogenic bacteria. The versatility of energy metabolism is better for C. violaceum than for bacteria from any other genus because the former generally has with six enzymes, whilst other bacteria lack one or more of these enzymes (Creczynski-Pasa et al., 2004). Metal cyanidation using a bacteria such as C. violaceum is believed to follow an indirect leaching mechanism involving two stages. First, glycine undergoes conversion to HCN (leaching chemical) by the associated enzymes under the metabolic activity of HCN synthase (stage I) (Knowles and Bunch, 1986). This is followed by reaction of the cyanide ions with the gold/copper substrate, leading to dissolution of metal (stage II), as presented in Figure 1. Actually, the amount of cyanide generated by C. violaceum strongly depends on the medium in which it Copyright © 2011 The Society of Chemical Engineers, Japan Fig. 1 Schematic of metal cyanidation using a microorganism is grown. In previous investigations, media with different organic components—peptone 1% (w/v), L-glutamate salts (Michaels and Corple, 1965); peptone 1% (wt/v), 2 mmol/L-glycine or 0.5 mmol/L-methionine (Rodgers and Knowles, 1978); and several other organic components (Collins et al., 1980) have been used. Besides organic components, some metal ions such as Na⫹ and Mg2⫹ added to the medium in low concentrations can catalyze and enhance the cyanide generation capacity of C. violaceum. Addition of Na2HPO4 in small amounts is also found to increase the amount of cyanide generated by such bacteria. In earlier researches, the effect of addition of these metal ions on the metabolic activity of bacteria has been studied using media containing different organic components such as tryptone (10 g/L) and yeast extract (5 g/L) (Faramarzi et al., 2004; Brandl et al., 2008), or polypeptone (10 g/L) and yeast extract (2 g/L) (Kita et al., 2005, 2008; Lawson et al., 1999). It is, therefore, very difficult to compare the effect of different metal ions on the cyanide generation capacity of C. violaceum in a particular medium. While the gold bioleaching process using a microorganism may be considered one of the most ecofriendly options, the dissolution rate may not be as fast as that in the case of direct addition of cyanide salts. Earlier researches (Kita et al., 2005; Lawson et al., 1999) have shown that the presence of some metal ions can improve the bacterial cyanidation process. An enhanced rate of gold dissolution was observed with cyanide solutions in presence of small amounts of Pb(NO3)2, which activated the gold surface, leading to higher reactivity and faster leaching (Sandenbergh and Miller, 2001; May et al., 2005). However, the effect of Pb2+ on the extent of cyanide generation by cyanogenic micro-organisms has not been investigated. For the application of cyanide-generating bacteria to gold and copper leaching, major emphasis has been placed on understanding the dissolution behavior of metal powders (Brandl et al., 2008; Kita et al., 2006; Smith and Hunt, 1985) in the presence of various microbes, and in a few cases the extraction of gold from ores (Lawson et al., 1999). Recently, efforts were made to treat secondary sources including waste printed circuit boards PCBs, by using cyanobacteria (Faramarzi et al., 2004; Brandl et al., 2008). In view of the fact that waste mobile phone PCBs are a rich source of gold, besides copper and other metals, the microbial recovery of gold by cyanogenic bacteria may be worthwhile to pursue as a part of the overall objective to develop a favorable recycling technology for waste PCBs. The residue from bioleaching could then be treated by a suitable hydrometallurgical process to extract copper and other metals. Thus, the effect of different metal ions including Pb2⫹ and Na2HPO4 on cyanide generation by C. violaceum in a medium with the predetermined organic components has been investigated in this study. The potential use of the biogenic cyanide produced from C. violaceum in the presence of these metal ions to dissolve gold from waste mobile phone PCBs has also been examined to establish an alternative recycling process. 1. Experimental 1.1 Microorganism and waste PCBs C. violaceum was supplied by Korea Collection for Type Cultures (KCTC), Biological Resource Center (BRC), Daejeon. Revival of C. violaceum in a medium comprising 5 g/L polypeptone and 3 g/L beef extract in 100 mL solution was performed at pH 6.8 and 30°C in an incubator shaker at 150 rpm for 2 d. To check the culture purity, the revived culture was incubated on an agar plate for 24 h at 37°C. Viable suspended cells were counted using the drop plate method with serial dilution in an agar medium (beef extract 3.0 g/L, peptone 5 g/L, agar 15 g/L). The plates containing pure cultures were sealed with parafilm and stored under refrigerator. A known volume of the culture medium containing viable cells was inoculated aseptically for investigating the effect of different metal ions on the amount of cyanide generated. In this research, a growth medium consisting of yeast extract (5 g/L), polypeptone (10 g/L), and glycine (5 g/L), designated as YP medium, was used. Samples of waste mobile phone PCBs for the bioleaching experiments were cut to 1 mm ⫻ 1 mm size by using scissors. Analysis showed a typical sample to consist of around 34.5 wt% copper and 0.025 wt% gold. The effect of different metal ions and Na2HPO4 on the cyanide production of C. violaceum in the YP medium was studied. Preliminary experiments were conducted by addition of NaCl, MgSO4, and FeSO4 to the cultures, in order to determine the metal ion concentrations at which the cell population was not significantly affected. By adding 0.17 mol/L NaCl, 4.0 ⫻ 10⫺3 mol/L MgSO4, 1.8 ⫻ 10⫺3 mol/L FeSO4, and a mixture of these salts, besides varying the Pb(NO3)2 concentration in the range 3.0 ⫻ 10⫺6 to 6.0 ⫻ 10⫺5 mol/L, the cyanide concentration in the inoculated medium was measured. The concentration of phosphate in the medium containing C. violaceum was varied in the range 1.0 ⫻ 10-2 to 3.0 ⫻ 10-2 mol/L by the addition of phosphate buffer. The metal content was analyzed by atomic absorption spectrometry (AAS, AAnalyst 400, Perkin Elmer Inc.). 693 1.2 Bioleaching All experiments were carried out in a 250-mL sterilized Erlenmeyer flask containing 200 mL of the YP medium. A 1 mL aliquot of C. violaceum under log phase culture conditions was added to the flask aseptically. The desired amounts of phosphate buffer solution and metal ions were then transferred to the flask. The pH of the solution was adjusted by the addition of 2.0 mol/L NaOH solution. The flask with the inoculated set was placed in an incubator shaker maintained at 30°C and incubated for a specified period. The supernatant sample was withdrawn every 24 h, and the cell count, pH, and quantity of cyanide generated were determined. For each inoculated set, a control experiment without metal ions was also carried out. No air supply was maintained during the experiments. The bacterial population was determined by counting the colonies, followed by cell count on an agar plate incubated at 37°C for 24 h, which was correlated with the optical density. The morphological features of C. violaceum were observed under a biological microscope (BX 51, Olympus Corp.) and the cell population was confirmed using a counting chamber. In order to examine the bio-dissolution of gold and copper from waste mobile phone PCBs, a known amount of the sample (1 mm ⫻ 1 mm in size) was placed in a 250-mL flask containing 200 mL of the medium inoculated with a 2% active culture of C. violaceum. The flasks were incubated at 30°C and 15 g/L pulp density in an orbital motion incubator with shaking at 150 rpm while maintaining the pH in the range 8.0–1.0. The supernatant was withdrawn at 24 h intervals to conduct a bacterial cell count, and to measure the pH, dissolved oxygen (DO), and cyanide concentration. After the bioleaching period was complete, the solution was filtered through Whatman 42 filter paper. The obtained clear solution was analyzed for gold and copper, and the percentage of metal leached was calculated (leaching efficiency, %). The residue obtained in each experiment was dissolved in aqua regia, and the metal in the leached and aqua regia solutions was analyzed by AAS to calculate the material balance. The concentration of cyanide generated by the bacteria during the process was determined colorimetrically by UV/Vis spectrometry (UV-1601PC, Shimadzu Corp.) at 580.5 nm using pyridine and barbituric acid (Clescert et al., 1998). For this, 5 mL of the sample supernatant was withdrawn from the experiment flask, and the pH was adjusted to 11.0 with 4 mol/L NaOH solution before filtration. To 10 mL of the clear solution, 4 mL of NaH2PO4 (138 g/L) was added, and the mixture was allowed to stand for 3 min; 2 mL of chloramine-T (10 g/L in distilled water) was added. After 5 min, 5 mL of the pyridine–barbituric acid mixture (75 mL pyridine, 15 g barbituric acid in 1 L water) was added to develop a stable violet color. The cyanide concentration was then analyzed colorimetrically at 580.5 nm against a standard solution. The DO value and pH were determined at 24-h intervals by a DO meter 694 (HQ40d, Hach Co.) and a pH meter (720A, Orion Research Inc.), respectively. Data given in the text are the average values of duplicate sets of experiments, with an accuracy of ⫾3.0%. 2. Results and Discussion 2.1 Cyanide generation by C. violaceum 2.1.1 Bacterial cell population and cyanide generation in the presence of metal ions The bacterial cell population was determined in presence of different metal ions at 30°C and an initial pH of 7.4. The results in Figure 2(a) show that the growth of C. violaceum reached the log phase after 2 d and that the maximum cell count was achieved in 5 d irrespective of the addition of metal ions. The decrease in the cell population after 5 d of incubation could be attributed to the decrease in cell multiplication once the optimum cell growth in the medium was attained, owing to the short stationary phase typically observed for cyano-bacteria. This also reflects the decreasing metabolic activity of such microbes with time (Knowles and Bunch, 1986). When no metal was present in the YP medium, the cell count reached a maximum of 12.3 ⫻ 108 cells/mL, with very little decrease in the bacterial population (11.3 ⫻ 108 cells/mL) until 7 d. Addition of 0.17 mol/L NaCl, 4.0 ⫻ 10⫺3 mol/L MgSO4, and 1.8 ⫻ 10⫺3 mol/L FeSO4 separately and the addition of a mixture of these salts to the medium adversely affected the cell population. The cell populations in the medium containing NaCl and the mixture of metal salts were much lower than those in the presence of MgSO4 and FeSO4. This could be attributed to the toxicity and inhibition of bacterial growth caused by the presence of Cl⫺ ions and the high concentration of metal ions. While Fe2⫹ was precipitated at pH 7.4 or higher, adsorption of a number of bacterial cells led to the observed decrease in the cell population in the presence of FeSO4. In contrast to the effect of metal ion concentrations on the cell population, the addition of a small amount of metal ions can increase cyanide generation by C. violaceum (Figure 2(b)). This is due to a catalytic effect enhancing the enzymatic process (Lawson et al., 1999). As can be seen in the figure, Fe2⫹ and Mg2⫹ were the most effective in enhancing cyanide generation, yielding cyanide concentrations of ⬃61 mg/L and 59 mg/L, respectively, in 5 d at an initial pH of 7.4. The cyanide concentration obtained after the addition of mixed metal ions was found to be approximately similar (⬃36 mg/L) to that observed in absence of these additives, after 7 d. Therefore, the YP medium with Mg2⫹ was chosen for the subsequent experiments. As air was not supplied to the medium, DO decreased quickly from 6.8 to 0.1 mg/L after the first day of incubation (Figure 2 (c)). However, this drop in the level of DO corresponded to an increase of the cell population as well as the cyanide concentration of the soluJOURNAL OF CHEMICAL ENGINEERING OF JAPAN Fig. 3 Effect of Pb(NO3)2 on cyanide generation by C. violaceum in the YP medium (4.0 ⫻ 10⫺3 mol/L MgSO4; temperature, 30°C; initial pH, 7.4). * YP media with 4.0 ⫻ 10⫺3 mol/L MgSO4 and 1.0 ⫻ 10⫺2 mol/L Na2HPO4 Fig. 2 Effect of metal ions on (a) cell population of C. violaceum, (b) cyanide generation, and (c) consumption of DO in the YP medium (temperature, 30°C; initial pH, 7.4) tion for up to 5 d after which a decreasing trend was observed. The consumption of dissolved oxygen is generally associated with the growth phase and bacterial respiration (Kita et al., 2008). The presence of various metal ions in the medium had no effect on the oxygen consumption during the bacterial growth. Thus, the influence of the dissolved oxygen concentration on cyanide generation by C. violaceum was not very significant after 24 h, and sustained VOL. 44 NO. 10 2011 bacterial metabolism was confirmed to improve the cyanide generation. During the growth of C. violaceum in the YP medium that was not supplemented with metal ions, the pH dropped quickly from 7.4 to 6.3 after one day and thereafter increased to 8.1 in 6 d. The presence of metal ions, particularly Mg2⫹, or a mixture of salts resulted in a similar trend with respect to the increase in pH, which was found to be nearly 8.5 after 7 d. The decrease in the pH of the medium was attributed to the acidification process for the conversion of organic compounds, particularly glycine, into carboxylic acids such as glyoxylic, cyanoformic, and oxamic acids. It was for this reason that the quantity of cyanide generated in first 2 d was not high (Figure 2(b)). The increase in pH after this point was accompanied by the increased generation of cyanide via the conversion of intermediate products such as carboxylic acids. Increased cyanide generation occurred from the second day onwards and reached the maximum after 5 d. A similar behavior has been observed previously (Smith and Hunt, 1985). In view of the improved dissolution of gold powder by C. violaceum in presence of Pb2⫹ (Sandenbergh and Miller, 2001; May et al., 2005), the effect of Pb(NO3)2 on cyanide generation in the YP medium was also investigated. Because of the small difference in the cyanide generation observed in the presence of Mg2⫹ and Fe2⫹, 4.0 ⫻ 10⫺3 mol/L of MgSO4 was added to the medium; the results are presented in Figure 3. It is apparent that the presence of Pb2⫹ in the medium affected the amount of cyanide produced by C. violaceum. With 3.0 ⫻ 10⫺6 mol/L (⬃1 mg/L) Pb(NO3)2 added to the YP medium, a higher quantity of cyanide was generated (66 mg/L in 5 d and 59 mg/L in 7 d). A Pb(NO3)2 concentration above 3.0 ⫻ 10⫺6 mol/L inhibited the bacterial cyanide generation. As observed below, the cyanide concentration was, however, found to be 695 Fig. 4 Cyanide generated by C. violaceum at varying phosphate concentrations in YP medium (4 ⫻ 10⫺3 mol/L MgSO4; temperature, 30°C; initial pH, 7.4) slightly higher (69 mg/L in 5 d) with 1.0 ⫻ 10⫺2 mol/L Na2HPO4 even in the absence of Pb(NO3)2. Incidentally, the presence of a similar amount of Pb(NO3)2 has been reported to improve gold recovery in the cyanidation (chemical) process (Sandenbergh and Miller, 2001). 2.1.2 Effect of Na2HPO4 Phosphate has been reported to increase the cyanogenesis process in C. violaceum (Rodgers and Knowles, 1978; Lawson et al., 1999). Its effect on cyanide generation in the YP medium was investigated in presence of 4.0 ⫻ 10⫺3 mol/L MgSO4. As can be seen from Figure 4, the addition of phosphate buffer affected the efficiency of cyanide generation. Cyanide generation was the highest (69 mg/L in 5 d) when 1.0 ⫻ 10⫺2 mol/L of Na2HPO4 was added to the medium; a further increase in the Na2HPO4 concentration decreased the cyanide concentration in the solution, as reported in previous observation (Lawson et al., 1999). The amount of cyanide generated (data not given in Figure 4) was found to increase to 73 mg/L at pH 9.0 in 5 d. The concentration was found to be 63 mg/L and 58 mg/L at pH 10.0 and 11.0, respectively, after the same culture period. 2.2 Bioleaching of gold and copper from waste mobile phone PCBs by C. violaceum Before examining the potential leaching of gold from waste mobile phone PCBs by C. violaceum in a YP medium containing 4.0 ⫻ 10⫺3 mol/L MgSO4, the effect of pH on cell population and cyanide generation was investigated. The results presented in Figure 5(a) show that the cell population decreased with an increase in the pH of the medium. At pH 7.4, the cell population reached a maximum (1.18 ⫻ 109 cells/mL) after 5 d of culture, while at pH 11.0, the cell population decreased to a lower value of 8.7 ⫻ 108 cells/mL. On the other hand, the cyanide concentration at 5 d, presented in Figure 5(b), increased from 59 mg/L to 68 mg/L when the pH was raised from 7.4 to 9.0. The cyanide concen696 Fig. 5 Effect of pH on cell population of C. violaceum (a) and cyanide generation (b) in the YP medium (4.0 ⫻ 10⫺3 mol/L MgSO4; temperature, 30°C) tration dropped to 56 mg/L at pH 9.0 after 7 d under these conditions. At pH 11.0, the cyanide concentration was low and was found to be 50 mg/L after 7 d. In the presence of the cyanide generated in the YP medium, both gold and copper can dissolve, forming complexes with the cyanide ions. Copper cyanide is produced in the form of various complexes, depending on the pH and other conditions, whereas gold mostly forms dicyanide anions [Au(CN)2⫺]. Since leaching of gold in the cyanidation process critically depends on the solubilization of copper in the sample, we have focused on the dissolution behavior of the latter along with that of in this research. As can be seen below (reactions 1–4), the dissolution of gold and copper to form cyanide complexes is thermodynamically favorable at 298 K in the presence of oxygen (Alonso-Gonzales et al., 2010; Rees and Vandeventer, 1999): 4 Au ⫹ 8CN⫺ ⫹ O 2 ⫹ 2 H 2 O ⫽ 4 Au ( CN )2⫺ ⫹ 4 OH⫺ (1) 0 ΔG298 ⫽⫺392.2 kJ/mol 4 Cu ⫹ 8CN⫺ ⫹ O 2 ⫹ 2 H 2 O ⫽ 4 Cu ( CN )2⫺ ⫹ 4 OH⫺ (2) 0 ΔG298 ⫽⫺322 kJ/mol JOURNAL OF CHEMICAL ENGINEERING OF JAPAN Fig. 6 Effect of pH on gold (a) and copper (b) leaching in the presence of MgSO4 in YP medium (4.0 ⫻ 10⫺3 mol/L MgSO4; pulp density, 15 g/L; temperature, 30°C) Cu ( CN )2⫺ ⫹ CN⫺ ⫹ O 2 ⫹ 2 H 2 O ⫽ Cu ( CN )32⫺ ⫹ 4 OH⫺ (3) 0 ΔG298 ⫽⫺228.6 kJ/mol Cu ( CN )3−2 ⫹ CN⫺ ⫹ O 2 ⫹ 2 H 2 O ⫽ Cu ( CN )4 3⫺ ⫹ 4 OH⫺ (4) 0 ΔG298 ⫽ ⫺163.3 kJ/mol Copper cyanide complexes, particularly tetra-cyanate, are more stable above pH 9.0 (Marsden and House, 1992) and tri-cyanate (stable in the pH range 7.0–12.0) can dissolve gold when the cyanide concentration is high (Vukcevic, 1996, 1997; Rees and Vandeventer, 1999). With low concentrations of free cyanide, the Cu(CN)2⫺ complex will be the most prevalent species, and it may not oxidize gold. 2.2.1 Effect of pH on cyanidation/leaching of metals in the presence of MgSO4 The effect of pH on the leaching of gold and copper from waste mobile phone PCBs was studied at 30°C and 15 g/L pulp density in presence of 4.0 ⫻ 10⫺3 mol/L MgSO4 in the YP medium; the results are shown in Figure 6. It is apparent that gold dissolution from the waste PCBs increased from 7.8% (0.35 mg/L) to a maximum VOL. 44 NO. 10 2011 value of 11% (0.46 mg/L) when the pH was increased from 8.0 to 11.0 (Figure 6 (a)). The trend was observed in the leaching of copper was found to be similar to that in the case of gold. The copper leaching efficiency was found to increase with pH, but a maximum of 11.4% (878 mg/L) was achieved at pH 10.0, and this value decreased marginally to 10.2% (815 mg/L) when the pH was further raised to 11.0 after 8 d of leaching (Figure 6(b)). Though, cyanide generation was higher (68 mg/L in 5 d and 56 mg/L in 7 d) at pH 9.0 (Figure 5(b)), high gold and copper leaching was observed at pH 11.0 and 10.0, respectively. This may be attributed to the requirement of a higher pH to form stable metal cyanide complexes under the present experimental conditions (Marsden and House, 1992). At pH 11.0, the cyanide concentration was analyzed to be 54 mg/L after 5 d and 50 mg/L after 7–8 d when gold leaching was the maximum (11%). 2.2.2 Effect of pH on cyanidation/leaching of metals in the presence of Na2HPO4 The effect of the addition of 1.0 ⫻ 10⫺2 mol/L Na2HPO4 to the YP medium on metal dissolution was investigated for varying pH values in the range 8.0–11.0; the results are presented in Figure 7. In the presence of Na2HPO4, the gold and copper dissolution trend was similar to that observed in the presence of Mg2+ up to a pH of 9.0. Gold leaching was not significantly different in the pH range 8.0–11.0, with a maximum dissolution of 9.2% gold at pH 11.0 (Figure 7(a)). Similarly, copper recovery was low within the pH range 8.0–9.0. However, copper recovery was found to increase substantially to 30.3% and 29.2% at higher pH values of 10.0 and 11.0 (Figure 7(b)), respectively. Thus, it is apparent that because the amount of cyanide generated (Figure 4) in the presence of Na2HPO4 is higher than that in the presence of MgSO4 (Figure 5(b)), copper leaching is more efficient than gold dissolution from PCBs under the above mentioned conditions. Therefore, pH is a critical factor affecting copper recovery, besides cyanide concentration. 2.2.3 Effect of Pb(NO3)2 on leaching/cyanidation of metals The results obtained for the addition of 3.0 ⫻ 10⫺6 mol/L Pb(NO3)2 to the YP medium in the absence of Na2HPO4 and MgSO4 are presented in Figure 8 for gold and copper bioleaching at pH 11.0 and 10.0, respectively. To compare the effect of other additives, the dissolution patterns of gold and copper in the presence of Na2HPO4 and MgSO4 are also shown for the same conditions. On the basis of the optimum pH derived from Figures 6 and 7, the leach recovery of gold and copper is also briefly mentioned (plots not presented) for pH 10.0 and 11.0, respectively, in the presence of a small amount of Pb(NO3)2. Copper bioleaching at pH 10.0 was found to be higher (38.1%) than that at pH 11.0 (29.3%, data not shown) in the presence of a small amount of Pb(NO3)2. However, higher gold bioleaching (10.1%) was observed at pH 11.0 (Figure 8(a)) than at pH 10.0 697 Fig. 7 Effect of pH on gold (a) and copper (b) leaching in the presence of Na2HPO4 in YP medium (1.0 ⫻ 10⫺2 mol/L Na2HPO4; pulp density, 15 g/L; temperature, 30°C) (9.4%, data not shown). Figure 8(b) also shows that copper bioleaching can be increased by 3 to 4 times when MgSO4 is substituted by Na2HPO4 and Pb(NO3)2 in the YP medium. As can be seen, copper bioleaching was 11.4% (878 mg/L) with MgSO4 which increased to 30.3% (2,156 mg/L) and 38.1% (2,668 mg/L) in the presence of small amounts of Na2HPO4 and Pb(NO3)2, respectively. In contrast, gold leaching in the presence of MgSO4 at pH 11.0 (11%) was only marginally higher than that in the presence of Na2HPO4 (9.2%) and Pb(NO3)2 (10.1%) (Figure 8(a)). From these results, it can be said that the addition of a low concentration of Na2HPO4 and Pb(NO3)2 enhances copper leaching because of the generation of a higher quantity of cyanide, (typically 69 mg/L and 66 mg/L, respectively) at pH 7.4, as compared to that in the presence of MgSO4 (59 mg/L cyanide at pH 7.4). Similarly, at higher pH values of 10.0 and 11.0, the concentration of cyanide generated in the presence of Na2HPO4 was clearly higher (63 mg/L and 58 mg/L in 5 d) than that in the presence of MgSO4 (57 mg/L and 54 mg/L cyanide), thus favoring copper bio-dissolution. 698 Fig. 8 Effect of different metal ions on the bioleaching of gold at pH 11.0 (a) and copper at pH 10.0 (b) in the YP medium (pulp density, 15 g/L; temperature, 30°C) The concentration of copper obtained in the presence of Pb(II) is higher than that obtained in the presence of Na2HPO4, probably because of the activation of the copper surface by Pb(II), which increases the reactivity of the surface toward cyanide. The higher solubility of copper over gold may also be related to the electrochemical behavior of the two metals, as gold is nobler 0 0 3⫹ (EAu /Au: 1.52 V) than copper (ECu2⫹/Cu: 0.34 V), and therefore, copper may dissolve preferentially while consuming bio-genic cyanide. Secondly, gold leaching will be facilitated in the presence of a high concentration of cyanide (Marsden and House, 1992), and the low level of bio-generated CN⫺ in the present case clearly favors the copper leaching at the expense of gold. As mentioned above, the Cu(CN)2⫺ complex available because of the low cyanide concentration and DO in this study will have very weak ability to dissolve gold. In Figure 9, the variation in the redox potential during the leaching of waste PCBs shows a low value of potential (about 2 mV at 6 d) at pH 11.0 (Figure 9(b)), favoring gold leaching over leaching at a low pH of 10.0 and below. The leaching behaviors of gold and copper at pH 9.0 and below were different from those at pH 10.0 and 11.0. This may be attributed to the stability of the JOURNAL OF CHEMICAL ENGINEERING OF JAPAN of Geoscience and Mineral Resources, 2010), by the physical beneficiation of waste PCBs, such as grinding and gravity separation, gold can be enriched (about 80–85 wt%) in the separated non-metallic portion containing a low quantities of metals. Therefore, it would be desirable to reduce the copper content by physical beneficiation and study the bioleaching of gold from the gold-enriched non-metallic component using C. violaceum. The major metallic portion enriched with copper and other metals can be treated by a suitable hydrometallurgical method to recycle the waste mobile phone PCBs completely. The main aspects of this research are as follows. The amounts of biogenic cyanide produced by C. violaceum in the presence of various metal ions in a particular medium (YP medium) were compared. The cyanide generated was used in gold leaching from waste mobile phone PCBs in order to recycle it, while understanding the effect of high copper content in the sample on the bio-dissolution of gold. Conclusion Fig. 9 Variation of the potential during bioleaching of copper at pH 10.0 (a) and gold at pH 11.0 (b) from waste mobile phone PCBs with C. violaceum in the YP medium (pulp density, 15 g/L; temperature, 30°C) iron hexacyanide complex [Fe(CN)64⫺] up to pH 9.0, which may decompose at higher pH and thus contribute to metal dissolution (Rees and Vandeventer, 1999). Our recent studies (data not included) involving the addition of 0.004%(v/v) hydrogen peroxide to supplement the DO level during the leaching of waste mobile phone PCBs by C. violaceum show that copper recovery and gold recovery can be improved by 8–10% and 2–3% over those obtained in this study, depending on the metal ions present. In view of the above results, it appears that gold bioleaching is adversely affected in the presence of a high copper concentration in the PCBs, and thus, it is necessary to reduce the copper content of the sample. The low leaching of gold may eventually affect the exploitation of the results. In order to make the present process/technology suitable for complete recycling of waste mobile phone PCBs, the gold recovery process must be improved. Precious metals, particularly gold, are mostly present as bonding wires and coated thin films in PCBs. As observed in our earlier research (Korea Institute VOL. 44 NO. 10 2011 Cyanide generation by cyanogenic bacteria— Chromobacterium violaceum in YP medium can be enhanced by adding a low amount of metal ions to the culture medium. The addition of MgSO4 and FeSO4 to the medium was found to be equally effective for cyanide generation by the bacteria, and the presence of Na2HPO4 and Pb(NO3)2 enhanced cyanide generation further. The dissolved oxygen concentration did not affect cyanide generation by C. violaceum. The results of bioleaching of valuable metals from waste mobile phone PCBs showed that the maximum amount of gold could be leached out (11% in 8 d) at pH 11.0 in the presence of 4.0 ⫻ 10⫺3 mol/L MgSO4 in the YP medium; however, the copper recovery was high (11.4%) at pH 10.0. When 1.0 ⫻ 10⫺2 mol/L Na2HPO4 and 3.0 ⫻ 10⫺6 mol/L Pb(NO3)2 were added to the YP medium, copper recovery was found to increase almost three fold (30.3%) and four fold (38.1%) at pH 10.0, respectively, in 8 d. The presence of phosphate and Pb(NO3)2 in the medium favored copper bio-dissolution, but it was not effective for gold leaching. The low concentration of cyanide generated by the metabolic activity of C. violaceum, and dissolved oxygen, favored copper bioleaching at the expense of gold from the PCBs. The results demonstrate the possibility of recovering gold and copper using C. violaceum. 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