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Preparation of Homogeneous Gold-Silver Alloy Nanoparticles Using the Apoferritin Cavity As a Nanoreactor Yongsoon Shin,*,† Alice Dohnalkova,‡ and Yuehe Lin† Pacific Northwest National Laboratory, 902 Battelle BouleVard, P.O. Box 999, Richland, Washington 99354 ReceiVed: NoVember 19, 2009; ReVised Manuscript ReceiVed: February 18, 2010
Homogeneous Au-Ag alloy nanoparticles have been synthesized in the cavity of horse spleen apoferritin (HSAF) by a diffusion technique. The Au-Ag nanoparticle cores are 5.6-6.3 nm in diameter with narrow size distribution (e1.0 nm), and their average diameter was gradually increased with an increase in the Ag content. The core formation ratios of Au-Ag-HASF samples are higher than 80%. These series of nanoparticles were applied for the reduction of 4-nitrophenol in the presence of NaBH4. As the Au content was increased in the Au-Ag-HSAF nanoparticles, the rate constant of the reduction was exponentially increased from 1.3 × 10-3 s-1 (pure Ag-HSAF) to 7.58 × 10-2 s-1 (pure Au-HSAF). These synthesized Au-Ag nanoparticles with different compositions will be further applicable in catalysis, sensing, and biomedical areas. 1. Introduction Metal alloy nanoparticles have attracted great attention because their changeable compositions lead to their unique sizedependent electronic, optical, and catalytic characteristics, which are different from those of the corresponding individual metal particles.1,2 For example, gold-containing bimetallic nanoparticles show enhanced catalytic activity.3 Ag-Au alloy nanoparticles are more catalytically active than monometallic nanoparticles, Ag or Au nanoparticles, in the oxidation of CO at low temperatures.4 Au alloys have been utilized as a Pt replacement for oxygen reduction in PEM fuel cells,5 and colloidal Au-Ag alloys have shown an enhanced electrocatalytic activity in direct borohydride fuel cells.6 In addition, the surface plasmon properties of Au-Ag alloy nanoparticles are continuously tunable because of the possibility of composition changes, whereas monometallic Ag and Au nanoparticles have relatively unchanging optical properties.7,8 Uniform Au-Ag alloy nanoparticles have been prepared via solution synthetic procedures.9 Some alloy nanoparticles have also been prepared by reduction with biological molecules.10 Numerous chemical methods have been employed for the synthesis of bimetallic nanoparticles using various reducing agents, including sodium borohydride,4 citrate,11 and hydrazine.12 One general method to prepare uniform metal nanoparticles is to use inverse micelle solution.13 This process requires high ratios of volume of toxic organic solvents or surfactants or block copolymers relative to the products. Recently, a biological method using protein cages as the biotemplate for the synthesis of metals, metal oxides, and semiconductors has been used.14 The delicate architecture and functional diversity of the biomolecules provide several important factors, such as sizes, shapes, arrangement, and compositions for the synthesis of inorganic nanoparticles.15 Since the protein cages are spatially organized, the obtained nanoparticles * To whom correspondence should be addressed. Phone: (509) 375-2693. Fax: (509) 375-2186. E-mail:
[email protected]. † Chemical and Materials Science Division, Pacific Northwest National Laboratory. ‡ Scientific Resources Division, Environmental Molecular Science Laboratory (EMSL).
SCHEME 1: Schematic Illustration of the Apoferritin-Templated Synthesis of Au-Ag-HSAF Nanoparticles
should be same in size and shape. Among cage-shaped proteins, horse spleen apoferritin (HSAF) is composed of 24 subunits, and the outer and inner diameters are 12 and 7 nm, respectively. Various kinds of metals, metal oxides, and semiconductors in the HSAF cavity have been synthesized;16 however, to our best knowledge, no report has been published on the synthesis of metal alloy nanoparticle inside the protein cage. There are two methods to synthesize metal nanoparticles using protein cages, diffusion and disassembly/reassembly methods. In the diffusion method, metal ions diffuse into the protein cage through 4 Å protein channels at pH 8.0-10.0, which connect the outside and inner cavity. In the disassembly/reassembly method, the subunits disassemble at low pH (2.0), and metalattached subunits reassemble into the original protein shape at higher pHs (8.0-10.0). The previous reports for the synthesis of Au17 and Ag18 in the HSAF cavity showed either broad particle size distributions or very low core formation ratios (CFRs) because they used simple water solvent, where metal precursors are not very stable. In this paper, we report the successful synthesis of homogeneous Au-Ag alloy nanoparticles in the HSAF cavity by the diffusion method (Scheme 1). Ag+ and AuCl4- ions competitively diffuse into the HSAF cavity, which is negatively charged, through the three-fold channels at pH 8.3. After dialysis against aqueous NH4OH solution to remove outer surface-bound metal
10.1021/jp911004a 2010 American Chemical Society Published on Web 03/04/2010
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Figure 1. (a) UV-vis spectra of Au-Ag-HSAF nanoaprticles with different compositions and (b) plot for surface Plasmon resonance as a function of Au-Ag composition. An arrow indicates an apoferritin peak at 283 nm.
ions, the metal ion mixture in the cavity is reduced upon adding NaBH4. Once a metal atom or metal ion mixture which acts as a nucleation center is formed, it acts as a catalyst for the reduction of the remaining metal ions present in the solution. The micro Au-Ag alloy particles produced at the initial stage of colloid formation subsequently agglomerate until they fully grow up to the HSAF cavity size. We prepared a series of Au-Ag-HSAF nanoparticles with different Au/Ag compositions and tested their catalytic activities on the reduction of 4-nitrophenol in the presence of NaBH4. 2. Experimental Section AgNO3, HAuCl4, and ammonium hydroxide were purchased from Sigma-Aldrich Chemical Co. and used without further purification. HSAF was purchased from Calzyme Laboratories, Inc. (San Luis Obispo, CA). A 10.0 mg portion of HSAF was dissolved in 15 mL of 7.5 mM NH4OH solution (pH 8.3), and the HSAF solution was divided into five 3.0 mL portions in 15 mL centrifuge tubes. Au-Ag alloy nanoparticles were synthesized by adding the different initial Au/Ag molar ratios (0:1, 0.25:0.75, 0.5:0.5, 0.75: 0.25, and 1:0) into the HSAF solutions. Different mole fractions of 0.01 M AgNO3 and 0.01 M HAuCl4 were added to each suspension. For example, 50 µL of 0.01 M AgNO3 and 50 µL of 0.01 M HAuCl4 were added to the HSAF solution for the 50/50 Au/Ag alloy system. After being incubated for 5 h, the solution mixtures were centrifuged at 10 000 rpm for 15 min in a 5804 R centrifuge (Eppendorf), followed by being passed through a PD-10 desalting column packed with Sephadex G-25 medium (1.5 cm × 5.5 cm, from Amersham Bioscience Corp.) against 7.5 mM NH4OH solution. HSAF was monitored by UV-vis spectroscopy at 280 nm. To each solution, 15 µL of 0.1 M NaBH4 was added. The solutions were immediately reduced and changed in color upon adding the NaBH4 solution. The solutions were centrifuged at 11 000 rpm, and the solution phases (alloy nanoparticle-HSAF products) were separated. The solutions were characterized using a HP 8453 UV-vis spectrophotometer. TEM images were obtained using a JEOL TEM 2010 microscope. The TEM sample was typically prepared by dropping sample suspension on a Cu grid coated with carbon films, and the TEM image of the products was obtained by staining with nano-W (Nanoprobes) solution on grids. The average particle sizes and the standard deviations were estimated from TEM images of over 250 particles using an image J program. The catalytic reduction of 4-nitrophenol by Au-Ag HSAF samples was conducted in the presence of NaBH4. In a standard quartz cell with 1.0 cm path length and about 3.0 mL volume,
0.2 mL of NaBH4 (0.1N) was mixed with 0.5 mL of 4-nitrophenol (0.2 mM). A 1.50 mL portion of DI water was then added. Immediately after addition of 0.3 mL of alloy samples, the UV-vis absorption spectra were recorded by a HP 8453 UV-vis spectrophotometer with 7 s time interval in a scanning range of 200-600 nm at 25 °C. 3. Results and Discussion Several buffer solutions, including Tris-HCl buffer and sodium phosphate buffer, were tried to stabilize Ag+ ion in the presence of AuCl4- ion at pH 8.0-10.0. It is important to completely dissolve Ag+ ions in the presence of Cl- from the AuCl4-, ensuring the reaction quotient (Q) is safely below the Ksp of AgCl(s), 1.6 × 10-10.19 Those buffers interacted with Ag+ and led to Ag precipitation. Therefore, low concentration NH4OH solution is suitable for Au-Ag alloy nanoparticle synthesis. Upon addition of Ag+ and AuCl4- ions into NH4OH solution (7.5 mM), the two metal ions are stabilized without any precipitation.20 Since the inner surface of HSAF is negatively charged by many amino acid residues,21 two metal precursors diffuse into the cavity until chemical equilibrium. Metal ion complexes outside the HSAF can be removed by dialysis against NH4OH solution. Finally, addition of unstable NaBH4 solution into the solutions reduces the metal ion mixtures to the corresponding alloy nanoparticles. Figure 1 shows the absorption spectra of the Au-Ag alloy nanoparticles, together with those of the pure forms. The absorption peaks arise from the surface Plasmon resonance (SPR). The SPR absorption peaks of pure Ag and Au nanoparticles are ∼402 and 524 nm, respectively. Those of Au-Ag alloy nanoparticles lie in the intermediate region, depending on the alloy compositions. The results confirm that the products are Au-Ag alloy nanoparticles rather than core-shell structures, where absorption peaks do not vary much as a function of composition. When the amount of gold ion in the solution compositions increases, the metal composition deviates from ideal behavior (solid straight line in Figure 1b). This might be a consequence of preferential chemical interaction between the silver complex and negatively charged cavity potential. TEM images of Au-Ag-HSAF nanoparticles negatively stained with nano-W staining confirmed that alloy dots weresurrounded by the protein shell (Figure 2).14c The cores are attributed to the Au-Ag materials due to nano-W stain’s being is too big to penetrate through the narrow channels. Au-AgHSAF nanoparticles are well-dispersed because the watersoluble HSAF shell prevents irreversible aggregation of the alloy particles and their precipitation. Pure alloy nanoparticles or aggregates are not observed outside HSAF, either. It should be
Homogeneous Au-Ag Alloy Nanoparticle Preparation
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Figure 2. TEM images of Au-Ag-HSAF nanoparticles with the negative stain method: (a) Au, (b) Au/Ag (3:1), (c) Au/Ag (1:1), (d), Au/Ag (1:3), (e) Ag, and (f) HR image of Au/Ag (3:1) with 2 nm a scale bar.
Figure 3. Histograms of the particle sizes of the samples: (a) Au, (b) Au/Ag (3:1), (c) Au/Ag (1:1), (d), Au/Ag (1:3), and (e) Ag. The size distribution of the particles was calculated from nonstained TEM images.
noted that Au-Ag alloy particles in HSAF-free control experiment resulted in the nonhomogeneous and bulk precipitation. Figure 3f shows a high-resolution TEM (HR-TEM) image of one core without staining. The core was a single crystalline lattice image. However, cores were usually polycrystalline due to their growth from multinucleation sites in the HSAF cavity. The size distribution of Au-Ag alloy cores measured by high resolution TEM and an image J program showed a narrow size distribution at ∼5.6-6.3 nm, and the average size distribution becomes a little broader and bigger as the Ag contents increase (Figure 3).22 The average diameters were (a) 5.6 ( 0.6, (b) 5.8 ( 0.8, (c) 6.0 ( 0.8, (d) 6.1 ( 0.7, and (e) 6.3 ( 1.0 nm. Usually, Au-Ag alloy nanoparticles prepared using a waterin-oil microemulsion or vacuum evaporation method showed
size distributions of 2-25 or 5-30 nm, respectively.6 This indicates that this method using the HSAF cavity can control the alloy particle size, and the size distribution is restricted by the cavity. Atomic compositions, average diameters, and absorbance peaks of Au-Ag-HSAF nanoparticles are summarized in Table 1. Atomic compositions estimated from energy dispersive X-ray spectroscopy (EDX, not shown here) are in good agreement with those of SPR of the samples. The CFRs of Au-AgHSAF nanoparticles,14b,c,16f,h which are measured from the TEM images and are represented as CFR(%) ) [Au-Ag-HSAF]/ [HSAF]total, were increased as an increase in the amount of silver ion in the solution composition (Figure 4).
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TABLE 1: Atomic Compositions, Average Diameters, and Absorbance Peaks of Au-Ag-HSAF Nanoparticles metal mole fraction
b
Ag
Au
1.00 0.75 0.50 0.25 0.00
0.00 0.25 0.50 0.75 1.00
atomic % of Aua av diam (nm)b λmax (nm) 0.00 ( 0.00 21.30 ( 2.80 40.89 ( 4.30 72.09 ( 2.70 100.00 ( 0.00
6.3 ( 1.0 6.1 ( 0.7 6.0 ( 0.8 5.8 ( 0.8 5.6 ( 0.6
402 425 441 494 524
a Atomic % of Au was estimated from EDX of nanoparticles. The particle size is averaged over 250 nanoparticles on TEM.
Figure 5. Successive UV-vis absorption spectra of the reduction of 4-nitrophenol by Au-Ag (3:1)-HSAF nanoparticles as a catalyst in the presence of NaBH4.
Figure 4. CFRs of Au-Ag-HSAF nanoparticles.
Au and Ag nanoparticles can be applied for the reduction of 4-nitrophenol into corresponding 4-aminophenol in the presence of NaBH4, which is one of the important applications of the metal nanoparticles.23,24 NaBH4 is not effective in this reduction unless provided with some catalysts to remove the kinetic barrier of the reaction to support electron relay for the reduction. Some catalysts, such as TiCl4,25 Au,21 and Ag22 have been used for this purpose. Here, five different Au-Ag-HSAF alloy nanoparticles, including pure Au and Ag samples, were investigated, for the first time, on the catalytic reduction of 4-nitrophenol in the presence of NaBH4. It has been observed that a red shift of the peak for 4-nitrophenol from 317 to 400 nm occurred immediately upon addition of NaBH4 due to the formation of 4-nitrophenolate ion in basic condition. The rapid catalytic conversion of the 4-nitrophenol to 4-aminophenol after addition of Au-Ag-HSAF samples was quantitatively monitored as a successive decrease in the peak height at 400 nm, corresponding to a change in solution color from light yellow to yellow-green. Figure 5 shows representative UV-vis spectra of the catalytic reduction of 4-nitrophenol into 4-aminophenol using the Au-Ag (3:1)-HSAF sample in the presence of NaBH4. Similar spectral
changes were also obtained for other Au-Ag-HSAF nanoparticle systems. The substrate reduction was not hindered as long as the concentration of NaBH4 remained constant during the reaction. In the absence of alloy catalysts, the absorption peak at 400 nm remained unaltered. This indicates that NaBH4 itself was not able to reduce 4-nitrophenolate ion directly. The absorption peak of 4-nitrophenolate ion at 400 nm decreases and disappears within a few minutes upon addition of Au-Ag-HSAF samples, with a concomitant blue shift of the peak from 400 to 290 nm, which indicates the formation of 4-aminophenol.26 Interestingly, we observed a concomitant peak at 494 nm during the successive decrease in the peak at 400 nm. The new peak represents Au-Ag (3:1) alloy. In the intermediate stage of the reduction, the peak due to the Au-Ag (3:1) plasmon band could not be observed and remained masked within the absorption band of the 4-nitrophenols, but finally, a clear absorption band at 494 nm due to the plasmon band appeared. In the catalytic reaction, a good linear fitting of ln(A400) versus the reaction time was obtained, indicating that pseudo-first-order kinetics could be used to calculate the kinetic rate constant. A good linear correlation, ln(A400) versus time, is obtained for all the systems studied. The kinetic rate constants of the Au-AgHSAF nanoparticles are estimated in Figure 6. The rate constant of Au-HSAF was 7.58 × 10-2 s-1 (7.0% standard deviation), which is significantly higher than that of other Au-Ag-HSAF nanoparticle catalysts, and that of Ag-HSAF was 1.3 × 10-3 s-1. When the amount of gold in the Au-Ag-HSAF samples
Figure 6. (a) Plots of ln(A400) against reaction time and (b) rate constants for the Au-Ag-HSAF nanoparticle catalytic reduction of 4-nitrophenol (7.0% standard deviation).
Homogeneous Au-Ag Alloy Nanoparticle Preparation linearly increases, the kinetic rate constants exponentially increase, indicating that Au could be much more efficient than Ag for the electron transfer from BH4- ion to 4-nitrophenol. To investigate the effect of the protein cage during the catalytic reduction process, Au nanoparticles (∼6.0 nm in diameter) were tested, and it was found that pure Au nanoparticles showed a rate constant (7.64 × 10-2 s-1) similar to that of Au-HSAF.21,23 4. Conclusions We have demonstrated the successful synthesis of Au-Ag nanoparticles in the HSAF cavity in an NH4OH solution by the diffusion method. The homogeneous 5.6-6.3 nm core size of Au-Ag-HSAF nanoparticles with over 80% CFRs showed strong surface plasmon resonances between 402 and 524 nm and a gradual increase in the particle diameter as of amount of Ag content increased. The Au-Ag-HSAF samples showed strong catalytic activity on the reduction of 4-nitrophenol in the presence of NaBH4. As the Au content increased in the Au-Ag-HSAF nanoparticles, the rate constants of the reduction exponentially increased from 1.3 × 10-3 s-1 to 7.58 × 10-2 s-1. This work may help the biomineralization study of multimetallic cores in the apoferritic cavity and fabrication of nanoelectronic devices with multicompositions. Acknowledgment. This work is supported by a laboratorydirected research and development program at Pacific Northwest National Laboratory (PNNL). This research was performed at the Environmental Molecular Sciences Laboratory (EMSL), a national scientific user facility sponsored by the U.S. Department of Energy’s Office of Biological and Environmental Research. PNNL is operated by Battelle for DOE under contract DEAC05-76RL01830. References and Notes (1) Liu, J. H.; Wang, A. Q.; Chi, Y. S.; Lin, H. P.; Mou, C. Y. J. Phys. Chem. B 2005, 109, 40–43. (2) Henglein, A. Chem. ReV. 1989, 89, 1861–1873. (3) Bond, G. C. Catal. Today 2002, 72, 5–9. (4) Wang, A. Q.; Liu, J. H.; Lin, S. D.; Lin, T. S.; Mou, C. Y. J. Catal. 2005, 233, 186–197. (5) Ferna´ndez, J. L.; Raghuveer, V.; Manthiram, A.; Bard, A. J. J. Am. Chem. Soc. 2005, 127, 13100–13101. (6) Atwan, M. H.; Northwood, D. O.; Gyenge, E. L. Int. J. Hydrogen Energy 2007, 32, 3116–3125. (7) Mallin, M. P.; Murphy, C. J. Nano. Lett. 2002, 2, 1235–1237. (8) Shin, Y.; Bae, I.-T.; Arey, B. W.; Exarhos, G. J. J. Phys. Chem. C 2008, 112, 4844–4848.
J. Phys. Chem. C, Vol. 114, No. 13, 2010 5989 (9) (a) Zheng, N.; Fan, J.; Stucky, G. D. J. Am. Chem. Soc. 2006, 128, 6550–6551. (b) Lu, X.; Tuan, H.-Y.; Chen, J.; Li, J.-Y.; Korgel, B. A.; Xia, Y. J. Am. Chem. Soc. 2007, 129, 1733–1742. (c) Wang, C.; Yin, H.; Chan, R.; Peng, S.; Dai, S.; Sun, S. Chem. Mater. 2009, 21, 433–435. (d) Wang, C.; Peng, S.; Chan, R.; Sun, S. Small 2009, 5, 567–570. (10) Senapati, S.; Ahmad, A.; Kahn, M. I.; Sastry, M.; Kumar, R. Small 2005, 1, 517–520. (11) Link, S.; Wang, Z. L.; El-Sayed, M. A. J. Phys. Chem. B 1999, 103, 3529–3533. (12) Chen, D.-H.; Chen, C.-J. J. Mater. Chem. 2002, 12, 1557–1562. (13) Murray, V. B.; Norris, D. J.; Bawendi, M. G. J. Am. Chem. Soc. 1993, 115, 8706–8715. (14) (a) Uchida, M.; Klem, M. T.; Allen, M.; Suci, P.; Flenniken, M.; Gillitzer, E.; Varpness, Z.; Liepold, L. O.; Young, M.; Douglas, T. AdV. Mater. 2007, 19, 1025–1042, and references therein. (b) Yoshimura, H. Colloids Surf., A 2006, 282, 464–470. (c) Yamashita, I. J. Mater. Chem. 2008, 18, 3813–3820. (d) Douglas, T.; Strable, E.; Willits, D.; Aitouchen, A.; Libera, M.; Young, M. AdV. Mater. 2002, 14, 415–418. (15) (a) Daniel, M. C.; Astruc, D. Chem. ReV. 2004, 104, 293–346. (b) Karz, E.; Willner, I. Angew. Chem., Int. Ed. 2004, 43, 6042–6108. (16) (a) Meldrum, C. F.; Wade, J. V.; Nimmo, L. D.; Heywood, R. B.; Mann, S. Nature 1991, 349, 684–687. (b) Mann, S. Nature 1993, 365, 499– 505. (c) Douglas, T.; Dickson, D. P. E.; Betterige, S.; Charnock, J.; Garner, C. D.; Mann, S. Science 1995, 269, 54–57. (d) Okuda, M.; Iwahori, K.; Yamashita, I.; Yoshimura, H. Biotech. Bioenger. 2003, 84, 187–194. (e) Li, M.; Viravaidya, C.; Mann, S. Small 2007, 3, 1477–1481. (f) Iwahori, K.; Yamashita, I. Nanotechnology 2008, 495601. (g) Liu, G.; Wu, H.; Dohnalkova, A.; Lin, Y. Anal. Chem. 2007, 79, 5614–5619. (h) Iwahori, K.; Yoshizawa, K.; Muraoka, M.; Yamashita, I. Inorg. Chem. 2005, 44, 6393–6400. (i) Ga´lvez, N.; Sa´nchez, P.; Domı´nguez-Vera, J. M.; SorianoPortillo, A.; Clemente-Leo´n, M.; Coronado, E. J. Mater. Chem. 2006, 16, 2757–2761. (j) Iwahori, K.; Yamashita, I. J. Cluster Sci. 2007, 18, 358– 370. (17) Zhang, L.; Swift, J.; Butts, C. A.; Yerubandi, V.; Dmochowski, I. J. J. Inorg. Biochem. 2007, 101, 1719–1729. (18) Domı´nguez-Vera, J. M.; Ga´lvez, N.; Sa´nchez, P.; Mota, A. J.; Trasobares, S.; Herna´ndez, J. C.; Calvino, J. J. Eur. J. Inorg. Chem. 2007, 4823–4826. (19) Jana, N. R.; Gearheart, L.; Murphy, C. J. J. Phys. Chem. B 2001, 105, 4065–4067. (20) (a) Smith, R. M.; Martell, A. E. Critical Stability Constants; Plenum Press: New York, 1975; Vol. 2, Amines. (b) Shriver, D. F.; Atkins, P.; Langford, C. H. Inorganic Chemistry, 2nd Ed.; Freeman: New York, 1994, pp 228-231. (21) Douglas, T.; Ripoll, R. D. Protein Sci. 1998, 7, 1083–1091. (22) Kim, M.-J.; Na, H.-J.; Lee, K. C.; Yoo, E. A.; Lee, M. J. Mater. Chem. 2003, 13, 1789–1792. (23) Liu, J.; Qin, G.; Raveendran, P.; Ikushima, Y. Chem.sEur. J. 2006, 12, 2131–2138. (24) Padhan, N.; Pal, A.; Pal, T. Colloids Surf., A 2002, 196, 247–257. (25) Kano, S.; Tanaka, Y.; Suzuki, S. Chem. Pharm. Bull. 1971, 19, 817–820. (26) (a) Hayakawa, K.; Yoshimura, T.; Esumi, K. Langmuir 2003, 19, 5517–5521. (b) Esumi, K.; Isono, R.; Yoshimura, T. Langmuir 2004, 20, 237–243.
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